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j.neurobiolaging.2018.07.014

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Neurobiology of Aging 71 (2018) 115e126
Contents lists available at ScienceDirect
Neurobiology of Aging
journal homepage: www.elsevier.com/locate/neuaging
Improved age-related deficits in cognitive performance and
affective-like behavior following acute, but not repeated,
8-OH-DPAT treatments in rats: regulation of hippocampal FADD
Elena Hernández-Hernández a, b, Antonio Miralles b, Susana Esteban b,
M. Julia García-Fuster a, *
a
University Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Balearic Islands Health Research Institute (IdISBa),
Palma, Spain
b
Laboratory of Neurophysiology, Department of Biology, University of the Balearic Islands, Palma, Spain
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 25 May 2018
Received in revised form 12 July 2018
Accepted 22 July 2018
Available online 29 July 2018
The aims of this study were (1) to behaviorally phenotype rats at different ages for both cognitive
performance and affect, (2) to evaluate the possible beneficial effects of 8-OH-DPAT (a 5-HT1A receptor
agonist) treatments on improving age-related behavioral deficits, and (3) to uncover putative key brain
targets (e.g., Fas-associated protein with death domain [FADD] and related partners) that might
contribute to the observed age-related behavioral changes. The principal results showed that acute, but
not repeated, 8-OH-DPAT treatments improved age-related deficits in cognitive performance and affect
while induced hypothermia. Moreover, multifunctional FADD protein decreased with age specifically in
the hippocampus (as compared to the prefrontal cortex) and was further decreased following acute 8OH-DPAT. The major conclusions indicate a parallelism between the beneficial effects observed
following acute 8-OH-DPAT on improving the negative consequences of aging on cognition and affect,
together with the acute induction of hypothermia and hippocampal FADD regulation. Because these
effects were not observed following repeated treatment (i.e., observed tolerance to acute hypothermia),
the results suggest 5-HT1A receptors desensitization and/or the activation of compensatory adaptive
mechanisms.
Ó 2018 Elsevier Inc. All rights reserved.
Keywords:
Aging
8-OH-DPAT
Hypothermia
Cognition
Hippocampus
FADD
1. Introduction
Aging is a complex, heterogeneous, and multifactorial process,
which is the consequence of multiple interactions between genes,
brain circuits, and environment (López-Otín et al., 2013; Marques
et al., 2010; Samson and Barnes, 2013) and is a strong risk factor
for most chronic disorders (e.g., Bektas et al., 2018). For example,
cognitive and memory capabilities are particularly vulnerable to the
aging process, both in humans and in laboratory rats (Mattson and
Magnus, 2006; Rosenzweig and Barnes, 2003). Moreover, aging has
been shown to increase the susceptibility to develop anhedonia
(i.e., the incapacity to experience pleasure, a key core symptom of
depression) in rats (Herrera-Pérez et al., 2008). In fact, depressionlike symptoms are associated with accelerated brain aging and are a
* Corresponding author at: IUNICS, University of the Balearic Islands, Cra. de
Valldemossa km 7.5, Palma E-07122, Spain. Tel.: þ34 971 259992; fax: þ34 971
259501.
E-mail address: j.garcia@uib.es (M.J. García-Fuster).
0197-4580/$ e see front matter Ó 2018 Elsevier Inc. All rights reserved.
https://doi.org/10.1016/j.neurobiolaging.2018.07.014
risk factor for different types of dementia (i.e., cognitive decline)
(Mckinney and Sibille, 2013).
During normal aging the most important age-related anatomical, physiological, and cognitive changes have been reported to
occur in neuronal circuits of the prefrontal cortex and hippocampus
of rodents and primates, which together indicated the existence of
age-related plasticity deficits (reviewed in Samson and Barnes,
2013). For example, the observed regression of terminal dendrites
and synaptic loss (with marked reduction of spine density in the
prefrontal cortex: Morrison and Baxter, 2012; Samson and Barnes,
2013) most probably contributes to the age-related decline in
cognition (learning and working memory) in the brain. In relation
to this, data from our research group recently showed that Fasassociated protein with death domain (FADD), a key cell fate
regulator balancing cell death and survival (e.g., García-Fuster et al.,
2007, 2008), was not only a putative biomarker of the cognitive
decline associated with the course of clinical dementia in an elderly
population (Ramos-Miguel et al., 2017), but it was also modulated
by procognitive drugs such as UK-14304 (i.e., a2-adrenoceptor full
agonist) in aged rats (see Hernández-Hernández et al., 2018).
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E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
Moreover, FADD was altered in major depression postmortem
brains (García-Fuster et al., 2014) and was regulated by antidepressant (García-Fuster and García-Sevilla, 2016) and monoaminergic drugs (i.e., including by 8-OH-DPAT, a 5-HT1A receptor
agonist; García-Fuster and García-Sevilla, 2015).
Interestingly, serotonin dysregulation is found to be involved in
many physiopathological cognitive processes, such as learning and
memory (Schmitt et al., 2006) and mood (Riedel et al., 2002). Plus,
antidepressant treatments that regulate the serotonergic system can
ameliorate the stress-related cognitive decline associated with pathological aging (e.g., Mowla et al., 2007; Wu et al., 2018). Among the
possible serotonergic targets, the activation of 5-HT1A receptors,
mainly somatodendritic autoreceptors, plays an important key
mechanism in cognitive functions in rats (e.g., Haider et al., 2012;
Wolff et al., 2004). In this regard, although some studies reported
improved cognitive performance following 8-OH-DPAT administration, others reported impaired results, an effect that seems to be
related to the dose administered: beneficial effects at lower doses (e.g.,
Haider et al., 2012; Inui et al., 2004), whereas larger or repeated doses
are associated with 5-HT1A receptor desensitization and postsynaptic
activation (Haider et al., 2012; Kreiss and Lucki, 1997). Moreover,
because 5-HT1A receptor dysfunction also plays an important role in
the mechanisms of depression (e.g., Kaufman et al., 2016), the
antidepressant-like effects of the agonist 8-OH-DPAT have been
strongly studied (Cervo and Samanin, 1987; Kennett et al., 1987).
Against this background, the aims of the present study were (1)
to behaviorally phenotype rats at different ages for both cognitive
performance and affective-like behavior, (2) to evaluate the
possible beneficial effects of 8-OH-DPAT (a 5-HT1A receptor agonist)
treatments on improving age-related behavioral deficits, and (3) to
uncover putative key brain targets (e.g., FADD and related partners)
that might contribute to the observed age-related behavioral
changes. Preliminary reports of this work were presented at the
30th European College of Neuropsychopharmacology Congress
(Hernández-Hernández and García-Fuster, 2017) and at the 37th
SEF National Meeting with guest society: the British Pharmacological Society (Hernández-Hernández et al., 2017).
2. Materials and methods
2.1. Animals
For this study, a total of 142 male Sprague-Dawley rats bred in
the animal facility at the University of the Balearic Islands were
used at 3 different age ranges: young adult rats (3e6 months old),
middle-aged rats (12e13 months old), and older rats
(16e18 months old). Rats were single housed in standard cages
under precise environmental conditions (22 C, 70% humidity, and
12 hours light/dark cycle, lights on at 8:00 AM) with ad libitum
access to a standard diet and tap water, and following the ARRIVE
guidelines (McGrath and Lilley, 2015) and according to standard
ethical guidelines (European Communities Council Directive 86/
609/EEC and Guidelines for the Care and Use of Mammals in
Neuroscience and Behavioral Research, National Research Council,
2003). The Local Bioethical Committee (UIB-CAIB) approved all
experimental procedures. All efforts were made to minimize the
number of rats used and their suffering. All procedures were performed during the light period (between 8:00 AM and 15:00 PM).
2.2. Experimental procedures
2.2.1. Study I: age-related changes in affective-like behavior and
cognitive performance
The first study aimed at evaluating possible physiological
changes in affective-like behavior and cognitive performance by
utilizing 27 rats at different ages (3, 6, and 13 months, n ¼ 9 rats
randomly allocated per age group of study; see Fig. 1A). All
behavioral data were analyzed by a blind experimenter to treatment group assignment. All behavioral equipment was cleaned in
between animals. Rats were handled for 2 days before any behavioral testing.
Changes in affective-like behavior were evaluated by 3 consecutive behavioral tests (i.e., spaced at least 2 days in between tests,
see Fig. 1A) that quantified core symptoms of depression (i.e.,
behavioral despair, anxiety-like behavior, and anhedonia). Behavioral despair was measured in the forced swim test (FST): following
a pretest training day (first day, 15 minutes of forced swimming in
an individual tankd41 cm high x 32 cm diameter, water to a depth
of 25 cm, and at 25 1 C), a test proceeds the next day in which
rats are videotaped while being forced to swim for 5 minutes (see
García-Cabrerizo et al., 2015 for further details). Videos were then
analyzed to obtain the time spent immobile versus active (mainly
climbing and swimming) (Behavioral Tracker software, CA, USA).
Anxiety-like behavior was evaluated by placing rats for 5 minutes in
a wall-enclosed circular open field (80 cm diameter). The test session was recorded to later evaluate the time spend in exploratorylike behavior (Walsh and Cummins, 1976). Then, anhedonia-like
behavior was evaluated by measuring the amount of sucrose
consumed by rats in a 2-bottle choice test (e.g., Slattery et al., 2007).
As described in more detail in a prior study from our group (see
García-Cabrerizo and García-Fuster, 2017), rats were weighted daily
throughout the procedure and were trained to drink from 2 bottles
placed on each side of the housing cage for 4 days. The first and last
days, both bottles were filled with water, and the other 2 days, 1
bottle contained 1% sucrose and the other one water. Bottles were
placed in alternate positions in the cage to avoid biased toward any
side and were weighted daily to calculate the amount of sucrose or
water consumed (g) per day per experimental group.
Following at least 3 days of rest after the last test, changes in
cognitive performance were evaluated in all rats by 2 consecutive
behavioral tests performed in different types of mazes that have been
developed to test hippocampal-dependent spatial learning (i.e.,
Barnes maze and 8-arm radial maze; e.g., Rosenzweig and Barnes,
2003; Vorhees and Williams, 2014). A period of 3 days of resting
was allowed in between tests (see Fig. 1A). The Barnes maze is a circular platform with 18 holes equidistantly located around the
perimeter. Below one of the holes there is a black “escape” box or
target. A bright light was used as a stimulus to find the target,
accentuating the natural agoraphobia of rats. Pretraining is performed
24 hours before training and testing day, and includes allowing the
animal to be for 10 seconds in the central box, for 180 seconds
exploring freely the maze, and for 120 seconds in the target box. The
next day, the pretest consists of 3 sessionsd10 minutes in between
trialsdin which the animal is allowed to be for 10 seconds in the
central box, for 180 seconds exploring freely the maze, and for
60 seconds in the target box. Finally, during the test the animal is
allowed to be for 10 seconds in the central box and 90 seconds of free
exploration. The amount of time spent, the number of errors
committed, as well as the strategy used to resolve the maze (i.e., direct,
serial, mixed), were used as a measure of spatial working memory
performance (e.g., Sarubbo et al., 2017).
For the next test and since the 8-arm radial maze (Panlab, S.L.,
Barcelona, Spain) requires motivation for food, rats were food
deprived for 36 hours before the beginning of the test. Briefly, and
as previously described (see Hernández-Hernández et al., 2018), the
maze consists of 8 equally spaced radial arms (50 cm long 12 cm
wide) with a food reward at the end of each arm and located in a
room with several external visual cues. On test day (see Fig. 1A), rats
were videotaped while allowed 20 minutes to resolve the maze.
Animal movement was monitored via a digital video tracking
E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
117
Fig. 1. Experimental design. (A) Study I: Age-related changes in affective-like behavior and cognitive performance. (B) Study II: Effects of 8-OH-DPAT treatments on cognitive
performance. (C) Study III: Effects of 8-OH-DPAT treatments on affective-like behavior. Abbreviations: D, day; FST, forced swim test; OF, open field; WB, Western blot.
system (LE 8300 with software SEDACOM v 1.3, Panlab, S.L., Barcelona, Spain). The amount of time spent (i.e., 8 arms visited before
the cutoff time of 20 minutes) and the number of correct responses
or errors committed (i.e., sum of reentry into arms and nonvisited
arms, see Hernández-Hernández et al., 2018) to complete the maze
was used as an indicative of spatial working memory performance.
2.2.2. Study II: effects of 8-OH-DPAT treatments on cognitive
performance
The second study aimed at evaluating possible beneficial effects
of treating rats with 8-OH-DPAT (i.e., acute vs. repeated treatments,
2 separate experimental procedures, see Fig. 1B) on ameliorating
age-related cognitive decay (i.e., Barnes maze). A blind experimenter to the treatment group assignment analyzed all behavioral
data. All behavioral equipment was cleaned in between animals.
Before any drug injection, and following 2 days of handling, rats
were pretrained in the Barnes maze as detailed above. The next day,
and for the acute experiment, a total of 60 rats were used and
treated with either 8-OH-DPAT (0.3 mg/kg, i.p.) or saline (0.9% NaCl,
1 mL/kg, i.p., control-treated rats) at 3 different ages (3, 12, and
18 months old; n ¼ 10 randomly allocated rats per age and group,
see Fig. 1B). Core body temperature was measured by a rectal probe
connected to a digital thermometer (Compact LCD display thermometer, SA880-1M, RS, Corby, UK) basally and 1 hour postinjection. Cognitive function was evaluated in the Barnes maze
1 hour postinjection as detailed above. On the other hand, a total of
36 rats were used for the repeated treatment paradigm and were
injected for 7 days with either 8-OH-DPAT (0.3 mg/kg, i.p.) or saline
(0.9% NaCl, 1 mL/kg, i.p., control-treated rats) at 3 different ages (3,
12, and 18 months old; n ¼ 6 randomly allocated rats per age and
group, see Fig. 1B). Core body temperature was measured on day 1,
2, and 6 of treatment basally and 1 hour postinjection. Cognitive
function was evaluated in the Barnes maze 24 hours after the last
injection (see Fig. 1B).
2.2.3. Study III: effects of 8-OH-DPAT treatments on affective-like
behavior
The third study aimed at evaluating possible beneficial effects of
treating rats with 8-OH-DPAT (i.e., acute vs. repeated treatments, 1
experimental proceduredrepeated measures, see Fig. 1C) on
ameliorating age-related deficits in affective-like behavior (i.e.,
behavioral despair measured in the FST). A blind experimenter to
the treatment group assignment analyzed all behavioral data. A
group of 19 rats with an average age of 16 months old were used for
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E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
this study. Rats were handled for 3 days before any behavioral
testing and were then exposed to the FST (as detailed before) for
15 minutes on pretest day and for 5 minutes on test day, in which
rats were videotaped and the videos were analyzed (Behavioral
Tracker software, CA, USA) to obtain baseline values of the time
spent immobile for each rat. Then, after 2 days of resting, groups of
rats (balanced by baseline immobility performance) were treated
with either 8-OH-DPAT (0.3 mg/kg, i.p., n ¼ 10) or saline (0.9% NaCl,
1 mL/kg, i.p., n ¼ 9, control-treated rats) (see Fig. 1C) and 1 hour
after treatment were reexposed to the FST (D1 of video analysis).
The next 6 days (i.e., for a total repeated treatment of 7 days), rats
received a daily injection of 8-OH-DPAT or saline. Then, 24 hours
after the last injection, rats were reexposed again to the FST (D8 of
video analysis).
2.3. Tissue collection
Rats were killed by decapitation at the indicated times: study I,
10 days after the last behavioral testing; study II, 1 hour postinjection for the acute treatment and 24 hours for the repeated
treatment (i.e., right after Barnes test; see Fig. 1). The hippocampus
and the prefrontal cortex (only evaluated for study I, see further
details in Results) were freshly dissected from the right half of the
brain, rapidly frozen on liquid nitrogen, and kept at 80 C until
further evaluations by Western blot experiments. To note that
brains from study III were not collected for neurochemical studies.
2.4. Immunoblots experiments for target protein quantification
Total homogenates of brain regions (the hippocampus and prefrontal cortex) were prepared with minor modifications as previously
described (García-Fuster et al., 2007). Each sample (40 mg of total
protein) was loaded in 10 % SDSePAGE minigels (Bio-Rad Laboratories, Hercules, CA, USA) that were resolved by electrophoresis and
then processed following standard immunoblotting procedures
(Hernández-Hernández et al., 2018). Membranes were incubated
overnight at 4 C in blocking solution containing the primary antibody anti-FADD (H-181) (dilution 1:5000; sc-5559, Santa Cruz
Biotechnology, CA, USA). Membranes were then incubated for 1 hour
in secondary horseradish peroxidase-linked anti-rabbit IgG antibody
(1:5000; Cell Signaling). Total FADD was immunodetected as a 51kDa dimeric form (see characterization in García-Fuster et al.,
2007) by exposing membranes, previously incubated with ECL reagents (Amersham, Buckinghamshire, UK), to an autoradiographic
film (Amersham ECL Hyperfilm) for 1e60 minutes. The amount of
total FADD for each sample was compared to that of control-treated
rats in each gel following quantification by densitometry (GS-800
Imaging Calibrated Densitometer, Bio-Rad). The quantification procedure for each sample was assessed, at least 3e5 times in different
gels, and the mean value was used as a final estimate. In all experiments, membranes were stripped and reincubated with b-actin
(1:10000; clone AC-15, Sigma-Aldrich, MO, USA), which was used as
a loading control.
Moreover, other cell fate markers related to multifunctional
FADD protein, such as p-FADD (dilution 1:750; # 2785, Cell
Signaling, MA, USA), antiapoptotic p-ERK1/2 (dilution 1:1000;
#9101L, Cell Signaling, MA, USA) (i.e., García-Fuster et al., 2007) or
proapoptotic p-JNK1/2 (Thr183/Tyr185) (dilution 1:2000; #9251L,
Cell Signaling, MA, USA) (i.e., Keller and García-Sevilla, 2015), and
the mitochondrial antiapoptotic Bcl-2 (DC21) (dilution 1:1000; sc783, Santa Cruz Biotechnology) (Boronat et al., 2001) or proapoptotic cytochrome c (dilution 1:5000; #3183977, BD Biosciences,
CA, USA) (Álvaro-Bartolomé et al., 2011), were also quantified in the
hippocampus of rats at different ages following the same experimental procedures as previously described.
2.5. Data and statistical analysis
All data were analyzed with GraphPad Prism, Version 6
(GraphPad Software, Inc, San Diego, CA, USA). Results are expressed
as mean values standard error of the mean (SEM). Study I: for the
FST, the open-field test, the sucrose preference test (i.e., 2-bottle
choiceddose of sucrose consumed in g of sucrose per kg of
weight), the Barnes maze (i.e., time and number of errors), the 8arm radial maze (i.e., time and number of correct responses) and
Western blot experiments, one-way analyses of variance (ANOVAs)
were used to ascertain differences among age groups (Dunnett’s
multiple post hoc comparison tests when appropriate). When
comparing the amount of water versus sucrose consumed (g) at
each age group of study, a two-way ANOVA with Day (D1 vs. D2,
first or last 24 hours of sucrose availability) and Bottle (water vs. 1%
sucrose) as independent variables was used (Sidak’s multiple post
hoc comparison test). Study II: the effects of 8-OH-DPAT (i.e., acute
or repeated) on body temperature, cognitive performance, or hippocampal FADD protein content were evaluated with two-way
ANOVAs, in which Treatment (control vs. 8-OH-DPAT) and either
Age (3, 12, and 18 months) or Day of treatment (D1, D2, and D6)
were treated as independent variables. Multiple t-tests were performed to compare the effect of treatment at each age of exposure.
Study III: the effects on the time spent immobile (sec) on the FST
were evaluated using two-way repeated-measures ANOVA, in
which Treatment (control vs. 8-OH-DPAT) and Day of behavioral
testing (baseline, D1: 1 hour following acute drug injection, D8:
24 hours following repeated drug treatment) were considered as
independent variables. Multiple t-tests were performed to compare
the effect of treatment at each day of analysis. The level of significance was set at p 0.05.
3. Results
3.1. Age-related deficits in affective-like behavior and cognitive
performance in rats
Age-related deficits in affective-like behavior were observed in
rats at different levels: (1) increased time spent immobile in the FST
(þ45 18 seconds, p < 0.05 when comparing 13 vs. 3 months old
following ANOVA: F2,24 ¼ 3.13, p ¼ 0.062, Fig. 2A), (2) decreased
time spent exploring in the OF (59 13 seconds, p < 0.001 when
comparing 13 vs. 3 months old following ANOVA: F2,24 ¼ 13.30, p <
0.001, Fig. 2B), and (3) decreased sucrose consumption in the 2bottle choice test (6 months: 0.78 0.25 g sucrose/kg;
13 months: 1.10 0.25 g sucrose/kg; p < 0.01 and p < 0.001 vs.
3 months old, respectively, following ANOVA: F2,24 ¼ 10.10, p <
0.001, Fig. 2C). To note that the accumulative drop in sucrose consumption (Fig. 2C) in aged rats was driven by the decline in consumption during the second day of exposure (6 months: 19 4 g
sucrose; 13 months: 26 4 g sucrose; p < 0.001 when comparing
D2 vs. D1 for both ages, Fig. 2C).
When evaluating age-related changes in cognitive performance
in rats, first with the Barnes maze and then with the 8-arm radial
maze (see Fig. 1 for schedule), the results for both tests were
different. In particular, no age-related changes were observed in the
Barnes maze as measured by the time needed to resolve the maze
(F2,24 ¼ 1.44, p ¼ 0.258) or the number of errors committed (F2,24 ¼
0.83, p ¼ 0.450, Fig. 2D). However, the same rats showed agedependent decreases in cognitive performance when tested in the
8-radial maze as measured by the time used (F2,24 ¼ 5.15, p ¼ 0.014)
and the number of correct responses (F2,24 ¼ 6.25, p ¼ 0.006,
Fig. 2E). In particular, 13-month-old rats needed more time (þ400 138 seconds, p < 0.05) and performed worse (2.22 0.67 correct
responses, p < 0.01) than 3-month-old rats (Fig. 2E). Interestingly,
E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
119
Fig. 2. Age-related deficits in affective-like behavior and cognitive performance in rats. (A) Time spent immobile (sec) in the forced swim test. (B) Time spent exploring (sec) in the
open field. (C) Sucrose consumption (g sucrose/kg) and sucrose versus water consumption (g) per day (D1: day 1 and D2: day 2) in the 2-bottle choice test (1% sucrose vs. water). (D)
Time spent (sec) and number of errors committed to complete the Barnes maze test. (E) Time spent (sec) and number of correct responses needed to complete the 8-arms radial
maze. Groups of analysis: 3, 6, and 13 months old rats, n ¼ 9 per age group. Columns represent mean SEM. *p < 0.05, **p< 0.01, ***p < 0.001 versus 3-month-old rats or Jp < 0.001
when comparing D2 versus D1 for water versus sucrose consumption. Abbreviations: D, day; SEM, standard error of the mean.
these data were complemented with another experiment performed in mice that also evaluated cognitive performance (i.e.,
sequential evaluation in the Barnes maze and then in the 8-arm
radial maze as above) at different ages (3, 8, 11, and 22 months
old, n ¼ 11, 9, 7, and 10, respectively, per age group). In particular,
and similarly to what was observed for rats, no age-related changes
were observed in the Barnes maze as measured by the time needed
to resolve the maze (F3,32 ¼ 0.35, p ¼ 0.790) or the number of errors
committed (F3,32 ¼ 0.33, p ¼ 0.807). However, the same mice
showed age-dependent decreases in cognitive performance when
tested in the 8-arm radial maze (F3,33 ¼ 3.81, p ¼ 0.019). Post hoc
comparisons revealed that middle age and older mice as compared
to 3-month-old mice needed more time to resolve the maze
(8 months: þ40 49 seconds, p > 0.05; 11 months: þ158 62 seconds, p < 0.05; 22 months: þ151 39 seconds, p < 0.05; data
not shown in Figures).
3.2. Age-related and region-specific decreases in FADD protein
content in rats
The results showed age-related decreases in FADD protein
content in the rat hippocampus (F2,24 ¼ 5.30, p ¼ 0.012). In
particular, rats at 6 and 13 months had less FADD protein content in
the hippocampus (26 8%, p < 0.001; 21 8%, p < 0.05,
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E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
respectively) as compared to 3-month-old rats (Fig. 3A). These experiments were complemented by analyzing in another set of rats
FADD protein regulation in the hippocampus with age (3, 12, and
18 months old, n ¼ 8 per age group). Thus, the results shown in
Fig. 3A were replicated by showing age-related decreases in FADD
(F2,21 ¼ 5.26, p ¼ 0.014)drats at 12 and 18 months had less FADD
protein content in the hippocampus (34 8%, p < 0.05; 34 9%,
p < 0.05, respectively) as compared to 3-month-old rats (data not
shown in Figures). Interestingly, these effects were region specific
as no significant changes were observed for FADD regulation in the
prefrontal cortex (F2,24 ¼ 0.67, p ¼ 0.520; Fig. 3B).
Besides total FADD, the regulation of its phosphorylated form was
also evaluated in the hippocampus during aging. The results showed
that the content of p-FADD protein did not change significantly with
age when compared with 3-month-old rats (see Table 1). These data
were again replicated with the complementary experiment previously described (F2,21 ¼ 2.25, p ¼ 0.131; 3 months: 101 10%;
12 months: 75 12%; 18 months: 69 12%; data not shown).
Moreover, none of the other cell fate markers that are related to
multifunctional FADD protein and that were evaluated in the hippocampus of rats at different ages were altered (see Table 1).
3.3. Hypothermic effects following acute and repeated 8-OH-DPAT
treatments in rats at different ages
When evaluating the acute effects of 8-OH-DPAT on core body
temperature, the results showed a significant effect of Treatment
(F1,40 ¼ 200, p < 0.001), but no effect of Age (F2,40 ¼ 0.30, p ¼ 0.739)
nor a Treatment Age interaction (F2,40 ¼ 0.52, p ¼ 0.597). Multiple
t-test comparisons revealed that acute 8-OH-DPAT induced significant decreases in core body temperature at all ages (3 months: 2.44
0.19 C; 12 months: 2.85 0.40 C; 18 months: 2.88 0.22 C;
at least p < 0.001 vs. control-treated rats, Fig. 4A).
Given that the effects of repeated 8-OH-DPAT treatments on
core body temperature varied with age (F2,15 ¼ 5.28, p ¼ 0.018),
each age was analyzed separately (see Fig. 4B). For 3-month-old
rats, the results showed a significant effect of Treatment (F1,10 ¼
17.57, p ¼ 0.002), but no effect of Day (F2,20 ¼ 1.05, p ¼ 0.370) nor a
Treatment Day interaction (F2,20 ¼ 0.30, p ¼ 0.745). Multiple t-test
Table 1
Hippocampal protein content in rats at different ages
Protein
3 mo
6 mo
p-FADD
p-ERK1/2
p-JNK1/2
Bcl-2
Cyt c
92 20
104 6
99 6
103 8
100 3
118
104
101
119
112
17
6
8
15
5
13 mo
ANOVA: FDFn,DFd, p value
76 14
106 3
95 4
96 13
113 4
F2,24
F2,23
F2,24
F2,24
F2,23
¼
¼
¼
¼
¼
1.48, p ¼ 0.248
0.03, p ¼ 0.966
0.24 p ¼ 0.792
0.86, p ¼ 0.435
2.69, p ¼ 0.089
Key: mo, months.
Data represent mean standard error of the mean of n experiments per group, and
expressed as percentage of the corresponding 3-month-old rats (100%). ANOVAs did
not detect significant changes at the different ages of study.
comparisons revealed that repeated 8-OH-DPAT induced in 3month-old rats significant decreases in core body temperature on
D1 (1.13 0.39 C; p < 0.05) and D2 (1.02 0.24 C, p < 0.05) but
not on D6 (0.77 0.35 C; p > 0.05) of treatment when compared
to control-treated rats (Fig. 4B). For 12-month-old rats, the results
showed a significant effect of Treatment (F1,9 ¼ 66.40, p < 0.001),
Day (F2,18 ¼ 3.57, p ¼ 0.049), and a Treatment Day interaction
(F2,18 ¼ 4.09, p ¼ 0.034). Multiple t-test comparisons revealed that
repeated 8-OH-DPAT induced in 12-month-old rats significant decreases in core body temperature on D1 (2.53 0.38 C; p < 0.05)
and D2 (2.01 0.38 C, p < 0.05) but not on D6 (0.89 0.29 C; p
> 0.05) of treatment when compared to control-treated rats
(Fig. 4B). Moreover, 8-OH-DPAT exerted significant differences in
body temperature on D6 as compared to D1 of treatment (p < 0.01,
see Fig. 4B), suggesting the induction of tolerance to the acuteinduced hypothermia. For 18-month-old rats, the results showed
a significant effect of Treatment (F1,9 ¼ 65.34, p < 0.001), but no
effect of Day (F2,18 ¼ 2.80, p ¼ 0.088) nor a Treatment Day
interaction (F2,18 ¼ 0.81, p ¼ 0.461). Multiple t-test comparisons
revealed that repeated 8-OH-DPAT induced in 18-month-old rats
significant decreases in core body temperature on D1 (1.50 0.16
C, p < 0.001), D2 (1.38 0.31 C, p < 0.001), and D6 (0.92 0.26
C, p < 0.05) of treatment when compared to control-treated rats
(Fig. 4B). At this age window of analysis, 8-OH-DPAT also exerted
significant differences in body temperature on D6 as compared to
D1 of treatment (p < 0.01, see Fig. 4B), which again suggested the
induction of tolerance to the acute-induced hypothermia.
Fig. 3. Age-related and brain region-specific decreases in FADD protein content in rats. (A) Hippocampus and (B) prefrontal cortex. Groups of analysis: 3, 6, and 13 months old rats,
n ¼ 9 per age group. Columns represent mean SEM of n experiments per group and expressed as a percentage of FADD protein content on the brain of 3-month-old rats. *p < 0.05
and **p < 0.01 versus 3-month-old rats. Bottom: representative immunoblots depicting labeling of FADD and the corresponding b-actin as a loading control. Abbreviation: SEM,
standard error of the mean.
E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
121
Fig. 4. Hypothermic effects following acute and repeated 8-OH-DPAT treatments in rats at different ages (3, 12, and 18 months old rats). (A) Effects of acute treatment with 8-OHDPAT (0.3 mg/kg, i.p.) or saline (0.9% NaCl, 1 mL/kg, i.p., control group) on rectal body temperature. Groups of treatment: 3, 12, and 18 months old rats, n ¼ 10 per age and treatment
group. Circles represent means SEM of the difference (D, 1 h, basal value) in body temperature ( C). ***p < 0.001 when compared with the corresponding age-matched control
group. (B) Effects of repeated 8-OH-DPAT treatment (0.3 mg/kg per day, i.p., 7 days) or saline (0.9% NaCl, 1 mg/kg per day, i.p., 7 days, control group) on rectal body temperature
(measured on day 1, 2, and 6 of treatment). Groups of treatment: 3, 12, and 18 months old rats, n ¼ 6 per age and treatment group. Circles represent means SEM of the difference
(D, 1h, basal value) in body temperature ( C). *p < 0.05, ***p < 0.001 when compared with the corresponding age-matched control group. Jp < 0.01 when compared with day 1 of
treatment. Abbreviation: SEM, standard error of the mean.
3.4. Procognitive-like effects following acute, but not repeated,
8-OH-DPAT treatments in rats at different ages
When analyzing the effects of acute 8-OH-DPAT treatment on
the time needed to complete the Barnes maze in rats at different
ages, a two-way ANOVA detected a significant effect of Age (F2,52 ¼
3.78, p ¼ 0.029) and Treatment (F1,52 ¼ 17.8, p < 0.001) but no Age Treatment interaction (F2,52 ¼ 0.63, p ¼ 0.539). Multiple t-test
comparisons revealed that acute 8-OH-DPAT reduced the time
spent to complete the maze in rats at all ages tested (3 months: 34
4 seconds, p < 0.05; 12 months: 41 2 seconds, p < 0.01;
18 months: 21 10 seconds, p > 0.05) as compared to controltreated rats (Fig. 5A). When analyzing the number of errors
committed to complete the maze, the main results showed an effect
of Treatment (F1,52 ¼ 15.7, p < 0.001), given the reduced number of
errors observed for rats at all ages (3 months: 4.8 0.8 number of
errors, p < 0.05; 12 months: 5.7 0.2 number of errors, p < 0.05;
18 months: 3.1 1.4 number of errors, p > 0.05) as compared to
control-treated rats (Fig. 5A). Moreover, the strategy used in
control-treated rats (n ¼ 10 per group) to resolve the maze varied
with age, fluctuating from a more serial/direct strategy (i.e., 8 rats
resolved the test with a serial and/or direct strategy at 3 months,
whereas at 12 months there were 6 rats and at 18 months there
were 4 rats) toward a mixed one at later ages (i.e., 2 rats resolved
the test with a mixed strategy at 3 months, whereas at 12 months
there were 4 rats and at 18 months there were 6 rats; Fig. 5A).
Interestingly, acute 8-OH-DPAT improved cognitive performance at
all ages by decreasing mixed strategies (i.e., at 3 months: from 2 rats
to 1; at 12 months: from 4 rats to 1; at 18 months: from 6 rats to 2)
while increasing either direct and/or serial strategies (see Fig. 5A). It
is worth mentioning that we were also aiming at evaluating the
effects of acute 8-OH-DPAT (0.3 and 1 mg/kg, i.p.) on the 8-arm
radial maze, but because the test requires motivation for food,
rats were food deprived and were too unresponsive to complete
cognitive testing (personal communication, data not shown in
Figures).
Contrarily to the acute effects, no improvement on cognitive
performance was observed following repeated 8-OH-DPAT. In
particular, a two-way ANOVA detected a significant effect of Age
(F2,27 ¼ 6.13, p ¼ 0.006) but no Treatment (F1,27 ¼ 2.64, p ¼ 0.116)
nor Age Treatment interaction (F2,27 ¼ 0.32, p ¼ 0.727) (Fig. 5B).
When analyzing the number of errors committed to complete the
maze, similar results were obtained: a significant effect of Age
(F2,27 ¼ 8.08, p ¼ 0.002) but no Treatment (F1,27 ¼ 0.69, p ¼ 0.414)
nor Age Treatment interaction (F2,27 ¼ 0.80, p ¼ 0.460) (Fig. 5B). It
is worth remarking the apparent improvement in cognitive performance (i.e., less time and errors to resolve the maze) observed in
rats at later ages (effect of Age describe above), as observed for 12
and 18 months old rats when compared to younger rats (3 months
old). Moreover, injecting rats daily for 7 days and exposing them to
the maze 24 hours later altered the strategy used to resolve the
maze when comparing it to the one followed by control-treated rats
exposed to the maze after only an acute injection (e.g., compare
control-treated groups from Fig. 5A vs. 5B). Following repeated
injections, all young control-treated rats (n ¼ 6) utilized a serial
strategy to resolve the maze, whereas at older ages the strategy
fluctuated toward a combination of direct, serial, and/or mixed. In
any case, repeated treatment with 8-OH-DPAT did not seem to alter
that pattern in a qualitative way (see Fig. 5B). Please note that when
evaluating the % number of rats that used each type of strategy and
122
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Fig. 5. Procognitive effects following acute, but not repeated, 8-OH-DPAT treatments in rats at different ages. (A) Effects of acute treatment with 8-OH-DPAT (0.3 mg/kg, i.p.) or
saline (0.9% NaCl, 1 mL/kg, i.p., control group) on the time spent (sec), the number of errors, and the strategy used to complete the Barnes maze. Groups of treatment: 3, 12, and
18 months old rats, n ¼ 10 per age and treatment group. Columns represent mean SEM. *p < 0.05 and **p < 0.01 when compared with the corresponding age-matched control
group. (B) Effects of repeated 8-OH-DPAT treatment (0.3 mg/kg per day, i.p., 7 days) or saline (0.9% NaCl, 1 mg/kg per day, i.p., 7 days, control group) on the time spent (sec), the
number of errors, and the strategy used to complete the Barnes maze. Groups of treatment: 3, 12, and 18 months old rats, n ¼ 6 per age and treatment group. Columns represent
mean SEM. Abbreviations: C, control; DPAT, 8-OH-DPAT; SEM, standard error of the mean.
due to the small sample size (n ¼ 6 rats per group) for this type of
analysis, the results are merely shown as a qualitative approximation and not as a quantitative outcome.
3.5. Antidepressant-like effects following acute, but not repeated,
8-OH-DPAT treatments in old rats
When analyzing the effects of acute versus repeated 8-OH-DPAT
treatments on affective-like behavior (i.e., behavioral despair
measured as the time spent immobile in the FST) in old rats (i.e.,
16 months old), a two-way repeated measures ANOVA detected a
significant Treatment Day of behavioral testing interaction
(F2,34 ¼ 16.30, p < 0.001) (Fig. 6). In particular, acute, but not
repeated, treatment with 8-OH-DPAT induced an antidepressantlike effect as measured by decreased time spent immobile in the
FST (D1, acute: 48 15%, p < 0.05; D8, repeated: þ18 10, p >
0.05) in aged rats (Fig. 6).
3.6. Acute, but not repeated, 8-OH-DPAT treatments further
decreases hippocampal FADD protein in rats at different ages
As shown in Fig. 7A, when evaluating the acute effects of 8-OHDPAT on FADD protein regulation in the hippocampus, a two-way
ANOVA detected a significant effect of Age (F2,54 ¼ 5.52, p ¼
0.007; expected as previously described in Fig. 3A) and Treatment
Fig. 6. Antidepressant-like effects following acute, but not repeated, 8-OH-DPAT
treatments in old rats. Time spent immobile (sec) in the forced swim test as measured
before any drug treatment (Baseline), on day 1 of treatment (acute effects, D1) and
24 hours following the last daily injection of the repeated treatment (D8). Groups of
treatment: control (0.9% saline, 1 mL/kg, i.p., n ¼ 9) and 8-OH-DPAT (0.3 mg/kg per day,
7 days, i.p., n ¼ 10). Circles represent means SEM. *p < 0.05 when compared with the
corresponding day-matched control group. Abbreviation: SEM, standard error of the
mean.
E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
123
Fig. 7. Acute, but not repeated, 8-OH-DPAT treatments further decreases hippocampal FADD protein in rats at different ages. (A) Effects of acute treatment with 8-OH-DPAT (0.3 mg/
kg, i.p.) or saline (0.9% NaCl, 1 mL/kg, i.p., control group) on FADD protein content in the hippocampus. Groups of treatment: 3, 12, and 18 months old rats, n ¼ 10 per age and
treatment group. Columns represent mean SEM of n experiments per group and expressed as a percentage of FADD protein content on the hippocampus of 3-month-old control
rats. *p < 0.05 and **p < 0.01 when compared with the corresponding age-matched control group. Bottom: representative immunoblots depicting labeling of FADD and the
corresponding b-actin as a loading control. (B) Effects of repeated 8-OH-DPAT treatment (0.3 mg/kg per day, i.p., 7 days) or saline (0.9% NaCl, 1 mg/kg per day, i.p., 7 days, control
group) on FADD protein content in the hippocampus. Groups of treatment: 3, 12, and 18 months old rats, n ¼ 6 per age and treatment group. Columns represent mean SEM of n
experiments per group and expressed as a percentage of FADD protein content on the hippocampus of 3-month-old control rats. Bottom: representative immunoblots depicting
labeling of FADD and the corresponding b-actin as a loading control. Abbreviations: C, control; DPAT, 8-OH-DPAT; SEM, standard error of the mean.
(F1,54 ¼ 23, p < 0.001) but not a Treatment Age interaction (F2,54 ¼
0.91, p ¼ 0.410). Multiple t-test comparisons revealed that acute 8OH-DPAT reduced FADD protein content in the hippocampus of rats
at different ages (3 months: 49 7%, p < 0.01; 12 months: 26 9%, p > 0.05; 18 months: 32 5%, p < 0.05) when compared to
control-treated rats (Fig. 7A).
Contrarily to the observed acute effects, no changes on hippocampal FADD protein content were observed following repeated 8OH-DPAT. In particular, a two-way ANOVA detected a significant
effect of Age (F2,29 ¼ 9.63, p < 0.001; expected as previously
described in Fig. 3A) but no effect of Treatment (F1,29 ¼ 0.25, p ¼
0.623) nor a Treatment Age interaction (F2,29 ¼ 0.51, p ¼ 0.606)
(Fig. 7B).
3.7. The hypothermic effects induced by acute 8-OH-DPAT are
associated with better cognitive scores and lower FADD content in
the hippocampus of rats at different ages
Pearson’s correlation coefficients were evaluated to test for
possible associations between changes in rectal temperature, the
time spent to resolve the Barnes maze, and the content of hippocampal FADD following acute or repeated 8-OH-DPAT treatments.
The main results showed that there was a positive correlation between the ability of 8-OH-DPAT to induce a hypothermic response
with an improved cognitive performance (i.e., time spent to resolve
the Barnes maze; r ¼ 0.414, n ¼ 58, p ¼ 0.001) and with decreased
FADD protein in the hippocampus (r ¼ 0.415, n ¼ 60, p ¼ 0.001)
following acute (Fig. 8A) but not repeated (Fig. 8B) administration
in rats at different ages. However, the time spent to resolve the
Barnes maze did not correlate with FADD protein content in the
hippocampus following acute (r ¼ 0.206, n ¼ 58, p ¼ 0.120; Fig. 8A)
or repeated (r ¼ 0.336, n ¼ 33, p ¼ 0.056; Fig. 8B) 8-OH-DPAT
treatments.
4. Discussion
The first part of the study showed age-related deficits in
affective-like behavior and cognitive performance in rats. In
particular, aging increased 3 core symptoms of depression (i.e.,
behavioral despair, anxiety-like behavior, and anhedonia) and
decreased cognitive performance (i.e., time spent and strategy used
to resolve the Barnes maze). Besides, multifunctional FADD also
decreased with age in rats, an effect that was specific to the
hippocampal region, as compared to the prefrontal cortex. Interestingly, the second part of the study proved that the observed agerelated deficits on cognitive performance and affect could be
reverted following acute, but not repeated treatments with 8-OHDPAT, a selective 5-HT1A receptor agonist. In addition, acute 8-OHDPAT further decreased (as compared to just the effect of aging) the
content of multifunctional FADD protein in the hippocampus.
Remarkably, there was a positive correlation between the ability of
acute 8-OH-DPAT to induce a hypothermic response in rats at
different ages with improved cognitive performance and FADD
protein downregulation. Overall, these results support a beneficial
effect of acute 8-OH-DPAT on improving the negative consequences
of aging.
When evaluating the effects of physiological aging on behavior,
and in particular age-related effects on negative affect, the present
data complement prior results (Herrera-Pérez et al., 2008; Mckinney
and Sibille, 2013) to demonstrate that depression-like symptoms are
present in the aged brain (i.e., effects observed at 13 months old),
including an earlier increased susceptibility to develop anhedonia (i.e.,
from 6 months on). In terms of how aging affects cognitive performance in rats, while prior data demonstrated age-related deficits in
the Barnes maze (see revision done by Rosenzweig and Barnes, 2003),
the present results showed no overall basal age differences in learning
acquisition as evaluated with this test. Notably, as stated in the results
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E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
Fig. 8. Association between variables. Scatter plot depicting a significant positive correlation between changes in core body temperature and the time needed to resolve the Barnes
maze (sec) and between changes in core body temperature and FADD protein content in the hippocampus following acute (A), but not repeated (B), 8-OH-DPAT treatments in rats at
different ages. No association was observed between the time needed to resolve the Barnes maze (sec) and FADD protein content in the hippocampus. Each circle represents an
individual rat. The solid line is the best fit for the correlations (r ¼ 0.414, n ¼ 58, ***p ¼ 0.001 and r ¼ 0.415, n ¼ 60, ***p ¼ 0.001). The dotted curves indicate the 95% confidence
interval for the regression line.
section, similar outcomes were also obtained for mice with aging, thus
suggesting that the discrepancy in overall performance in the Barnes
maze among different research groups might be caused by dissimilarities in behavioral settings. In any case, it is worth remarking that
although no overall changes were observed in cognitive performance
in the Barnes test, the strategy used to resolve the maze, both for rats
and mice, worsen with age, fluctuating from a more serial/direct
strategy toward a mixed one at older ages, which is also an indicative
of cognitive dysfunction (e.g., Harrison et al., 2018). In this line of
thought, when spatial working memory was evaluated in the 8-arm
radial maze, rats (and mice) showed impaired performance with age
(i.e., effects observed at 13 months old), similarly to prior published
data (Rosenzweig and Barnes, 2003).
The neurochemical results showed region-specific decreases in
FADD protein content with age (i.e., from 6 months on) in the
hippocampus, as compared to the prefrontal cortex. Although both
regions play a crucial role in processing spatial information (e.g.,
Samson and Barnes, 2013), they subserve different components of
working memory in rats (Yoon et al., 2008). Because the hippocampus of aged rats sustains a loss of synapses in the dentate gyrus,
these age-related deficits in plasticity (i.e., FADD) may affect
cognition by changing the dynamics of the hippocampal network
(e.g., Rosenzweig and Barnes, 2003). Although FADD was initially
described as a proapoptotic marker (Chinnaiyan et al., 1995), over
the years it has proven to be a key marker of neural plasticity (e.g.,
Alappat et al., 2005; García-Fuster et al., 2016) that can be regulated
as an adaptive response to a prior insult (i.e., negative effects of
aging). In fact, this idea is supported by recent data from our
research group that associated loss of FADD with aging as evaluated
in the postmortem human brain of an elderly population (RamosMiguel et al., 2017). Another study reported enhanced FADD
phosphorylation (p-FADD) as a primary cause of reactive oxygen
species (ROS) accumulation during aging (Cheng et al., 2014). Thus,
considering that FADD and p-FADD forms normally regulate in an
opposite manner (e.g., García-Fuster and García-Sevilla, 2015, 2016;
García-Fuster et al., 2016), enhanced levels of p-FADD during aging
(Cheng et al., 2014) would support FADD downregulation. However,
no changes were detected in the hippocampus for p-FADD form or
for any of the other evaluated targets (e.g., p-ERK1/2, p-JNK1/2, Bcl2, and cytochrome c), thus suggesting a more prominent role for
FADD regulation during aging. In fact, lower FADD (but not p-FADD)
content in the postmortem human brains was associated with
E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
lower cognitive function as obtained by antemortem clinical diagnoses (see more details in Ramos-Miguel et al., 2017). In summary, so far, the present results validate the use of laboratory rats as
a good preclinical model to mimic the negative impact of aging
observed in humans (i.e., behavioral and hippocampal dysfunction),
and therefore a great tool to explore possible beneficial treatment
options and putative molecular correlates.
Therefore, and against these results, the second part of the
study evaluated the possible beneficial effects of 8-OH-DPAT (a 5HT1A receptor agonist) treatments on improving age-related
behavioral deficits in cognition and affect. As expected, 8-OHDPAT induced an acute hypothermic response, which was
observed 60 minutes postinjection (see Yu and Lewander, 1997)
and had similar effects on core body temperature for all ages
evaluated (see similar results, Darmani and Ahmad, 1999; Robson
et al., 1993). Following repeated 8-OH-DPAT, the results showed
the induction of tolerance to the acute-induced hypothermia (e.g.,
Johansson-Wallsten and Meyerson, 1994), which could be the
result of 5-HT1A receptor desensitization (e.g., Bouaziz et al., 2014;
Kreiss and Lucki, 1997). Similarly, acute, but not repeated 8-OHDPAT improved cognitive performance at different ages, by
decreasing the overall time spent and the number of errors
committed, and by tentatively improving the strategy used to
resolve the Barnes maze (i.e., toward a more direct one) (see also
Harrison et al., 2018). Therefore, our results complement prior
data supporting the beneficial procognitive effects of acute 8-OHDPAT administered at low doses (e.g., Haider et al., 2012; Inui et al.,
2004) and the lack of effects following repeated doses (i.e., probably associated with 5-HT1A receptor desensitization and postsynaptic activation; Haider et al., 2012; Kreiss and Lucki, 1997).
More importantly, our results further extend this prior knowledge
by proving that 8-OH-DPAT beneficial effects are also observed at
different ages, with lower overall impact on older ages (18 vs. 12 or
3 months of age). Moreover, even though antidepressant efficacy
seems to be reduced in elderly patients with depression, acute 8OH-DPAT was capable of inducing a potent antidepressant-like
effect in 16-month-old rats in the FST, thus pointing at this drug
as a good acute treatment for the behavioral dysfunctions associated with aging. Interestingly, following repeated 8-OH-DPAT
administration the antidepressant-like effect was dissipated. It is
well known that the effects of 8-OH-DPAT are mainly mediated by
5-HT1A receptors, which are confined to the somatodendritic
compartment of both serotonergic neurons and target neurons of
serotonergic projections where they act as autoreceptors and
heteroreceptors, respectively. Moreover, repeated 5-HT1A receptor
stimulation induced autoreceptor desensitization at different
rates in different brain regions (e.g., Kreiss and Lucki, 1997). In fact,
agonist-induced internalization of 5-HT1A receptors, which depends on agonist efficacy and neuronal phenotype, might underlie
receptor desensitization following activation (Bouaziz et al., 2014).
This adaptive compensatory response following repeated 8-OHDPAT treatments might help explain the lack of effects observed in
the present study (i.e., behavioral effects as well as the induction
of tolerance to the acute-induced hypothermia). In the same line
of thought, the content of multifunctional FADD protein, which
was decreased with aging, was further decreased following acute,
but not repeated, 8-OH-DPAT in the hippocampus of rats at
different ages. The acute downregulation of brain FADD by 8-OHDPAT, and the involvement of 5-HT1A receptors in this modulation
(i.e., pharmacological antagonism with WAY-100635), was previously evaluated by our research group in young adult rats (GarcíaFuster and García-Sevilla, 2015). The present data replicated that
study to also extend 8-OH-DPAT effects on FADD at later age
windows in life. Thus, brain FADD content is decreased with aging
(present results for rodents; see Ramos-Miguel et al., 2017 for
125
human studies and for further discussion of the implications of
such downregulation, e.g., adaptive response to a prior insult and/
or synaptic dysfunction or dysplasticity), lower FADD content
correlated with a poorer cognitive performance and odds rate of
clinical dementia (Ramos-Miguel et al., 2017), and drugs that
regulate FADD could be expected to improve cognitive performance (e.g., Hernández-Hernández et al., 2018). Therefore, the
present data show that acute 8-OH-DPAT improved cognitive
performance while further decreased hippocampal FADD. The
precise mechanism that could explain how FADD downregulation
could participate in the potential beneficial effects mediated by
acute 8-OH-DPAT deserves further studies (see also García-Fuster
and García-Sevilla, 2015 for results suggesting a role of decreased
FADD in neuroplastic/neuroprotecive mechanisms). In fact, the
neuroprotective effects of acute 8-OH-DPAT (i.e., improved
cognition, FADD regulation) were associated with the induction of
hypothermia in rats of different ages. Although small drops in
temperature are normally used in the clinic to improve the
neurological outcome under various pathological conditions (e.g.,
stroke, brain injury; see e.g., revision in Kurisu and Yenari, 2018),
the specific mechanisms of hypothermic neuroprotection remain
unclear. Interestingly and in line with the current observed
decrease in FADD protein content, a prior study suggested that 8OH-DPAT reduced brain tissue injury in rats by diminishing neural
cell apoptosis (e.g., decreased caspase-3 and Bax protein expression), effects that were observed in association with mild hypothermia (Mao et al., 2013). Thus, how FADD is being regulated and
the role that might be playing in the hypothermic response and/or
the procognitive effects induced by 8-OH-DPAT deserves future
studies. Interestingly, FADD dysregulation during aging (and its
regulation by 8-OH-DPAT treatments) adds to the age-related
deficits that regulate hippocampal dynamics and might induce
behavioral dysfunctions (e.g., Rosenzweig and Barnes, 2003).
The major conclusions of the present study indicate a parallelism between the beneficial effects observed following acute, but
not repeated, 8-OH-DPAT on improving the negative consequences
of aging on cognition and affect, together with the induction of
hypothermia and hippocampal FADD regulation. Because these effects were not observed following repeated treatments, the results
suggest desensitization of 5-HT1A receptors and/or activation of
compensatory adaptive mechanisms.
Disclosure statement
The authors have no actual or potential conflicts of interest to
disclose.
Acknowledgements
This work was supported by the following funding sources:
SAF2014-55903-R (Ministerio de Economía, Industria y Competitividad, MINEICO) and AAEE098/2017 (Consejería de Innovación, Investigación y Turismo del Gobierno de las Islas Baleares,
CAIB y del Fondo Social Europeo, FEDER) to M.J.G-F. The authors
thank Antonio Crespo for skillful technical assistance.
References
Alappat, E., Feig, C., Boyerinas, B., Volkland, J., Samuels, M., Murmann, A.E.,
Thorburn, A., Kidd, V.J., Slaughter, C.A., Osborn, S.L., Winoto, A., Tang, W.J.,
Peter, M.E., 2005. Phosphorylation of FADD at serine 194 by CKIalpha regulates
its nonapoptotic activities. Mol. Cell 19, 321e332.
Álvaro-Bartolomé, La Harpe, R., Callado, L.F., Meana, J.J., García-Sevilla, J.A., 2011.
Molecular adaptations of apoptotic pathways and signaling in the cerebral
cortex of human cocaine addicts and cocaine-treated rats. Neuroscience 196,
1e15.
126
E. Hernández-Hernández et al. / Neurobiology of Aging 71 (2018) 115e126
Bektas, A., Schurman, S.H., Sen, R., Ferrucci, L., 2018. Aging, inflamation and the
environment. Exp. Gerontol. 105, 10e18.
Boronat, M.A., García-Fuster, M.J., García-Sevilla, J.A., 2001. Chronic morphine induces
up-regulation of the pro-apoptotic Fas receptor and down-regulation of the antiapoptotic Bcl-2 oncoprotein in rat brain. Br. J. Pharmacol. 134, 1263e1270.
Bouaziz, E., Emerit, M.B., Vodjdani, G., Gautheron, V., Hamon, M., Darmoon, M.,
Masson, J., 2014. Neuronal phenotype dependency of agonist-induced internalization of the 5-HT1A serotonin receptor. J. Neurosci. 34, 282e294.
Cervo, L., Samanin, R., 1987. Potential antidepressant properties of 8-hydroxy-2-(din-propylamino) tetrain, a selective serotonin1A receptor agonist. Eur. J.Pharmacol. 144, 223e229.
Cheng, W., Zhang, R., Yao, C., He, L., Jia, K., Yang, B., Du, P., Zhuang, H., Chen, J., Liu, Z.,
Ding, X., Hua, Z., 2014. A critical role of Fas-associated protein with death
domain phosphorylation in intracellular reactive oxygen species homeostasis
and aging. Antioxid. Redox Signal. 21, 33e45.
Chinnaiyan, A.M., O’Rourke, K., Tewari, M., Dixit, T.V., 1995. FADD, a novel death
domain-containing protein, interacts with the death domain of fas and initiates
apoptosis. Cell 81, 505e512.
Darmani, N.A., Ahmad, B., 1999. Long-term sequential determination of behavioral
ontogeny of 5-HT1A and 5-HT2 receptor functions in the rat. J. Pharmacol. Exp.
Ther. 288, 247e253.
García-Cabrerizo, R., Keller, B., García-Fuster, M.J., 2015. Hippocampal cell fate
regulation by chronic cocaine during periods of adolescent vulnerability: consequences of cocaine exposure during adolescence on behavioral despair in
adulthood. Neuroscience 304, 302e315.
García-Cabrerizo, R., García-Fuster, M.J., 2017. Methamphetamine binge administration dose-dependently enhanced negative affect and voluntary drug consumption in rats following prolonged withdrawal: role of hippocampal FADD.
Addict. Biol. https://doi.org/10.1111/adb-12593.
García-Fuster, M.J., Miralles, A., García-Sevilla, J.A., 2007. Effects of opiate drugs on
Fas- associated protein with death domain (FADD) and effector caspases in the
rat brain: regulation by the ERK1/2 MAP kinase pathway. Neuropsychopharmacology 32, 399e411.
García-Fuster, M.J., Ramos-Miguel, A., Miralles, A., García-Sevilla, J.A., 2008. Opioid
receptor agonists enhance the phosphorylation state of Fas-associated death
domain (FADD) protein in the rat brain: Functional interactions with casein
kinase I-alpha, G-alpha-i proteins, and ERK1/2 signaling. Neuropharmacology
55, 886e899.
García-Fuster, M.J., Díez-Alarcia, R., Ferrer-Alcón, M., La Harpe, R., Meana, J.J., GarcíaSevilla, J.A., 2014. FADD adaptor and PEA-15/ERK1/2 partners in major depression
and schizophrenia postmortem brains: basal contents and effects of psychotropic
treatments. Neuroscience 277, 541e551.
García-Fuster, M.J., Álvaro-Bartolomé, M., García-Sevilla, J.A., 2016. The Fas receptor/
Fas-associated protein and cocaine. In: Preedy, V.R. (Ed.), Neuropathology of
Drug Addictions and Substance Misuse, Volume 2, Chapter 6. Academic Press
(Elsevier), pp. 63e73 eBook ISBN: 9780128003756. Hardcover ISBN:
9780128002124.
García-Fuster, M.J., García-Sevilla, J.A., 2015. Monoamine receptor agonists, acting
preferentially at presynaptic autoreceptors and heteroreceptors, downregulate
the cell fate adaptor FADD in rat brain cortex. Neuropharmacology 89, 204e214.
García-Fuster, M.J., García-Sevilla, J.A., 2016. Effects of anti-depressant treatments
on FADD and protein in rat brain cortex: enhanced anti-apoptotic p-FADD/FADD
ratio after chronic desipramine and fluoxetine administration. Psychopharmacology 233, 2955e2971.
Haider, S., Khaliq, S., Tabassum, S., Haleem, D.J., 2012. Role of somatodendritic and
postsynaptic 5-HT1A receptors on learning and memory functions in rats.
Neurochem. Res. 37, 2161e2166.
Harrison, F., Reisere, R., Tomarken, A., Mcdonald, M., 2018. Spatial and nonspatial
strategies in the Barnes maze. Learn Mem. 13, 809e819.
Hernández-Hernández, E., Miralles, A., Esteban, S., García-Fuster, M.J., 2018.
Repeated treatment with the alpha-2-adrenoceptor agonist UK-14304 improves
cognitive performance in middle-age rats: role of hippocampal Fas-associated
death domain. J. Psychopharmacol. 32, 248e255.
Hernández-Hernández, E., García-Fuster, M.J., 2017. Aged-related changes in negative affect and cognitive performance in Sprague-Dawley rats: possible role of
the cell fate regulator FADD. Eur. Neuropsychopharmacol. 27, S680eS681.
Hernández-Hernández, E., Miralles, A., Esteban, S., García-Fuster, M.J., 2017. Acute
treatment with 8-OH-DPAT improves cognition in rats by changing the strategy
used to resolve a maze at different ages. Basic Clin. Pharmacol. Toxicol. 121
(Suppl. 2), 44.
Herrera-Pérez, J.J., Martínez-Mota, L., Fernández-Guasti, A., 2008. Aging impairs the
antidepressant-like response to citalopram in male rats. Eur. J. Pharmacol. 633,
39e43.
Inui, K., Egashira, N., Mishima, K., Yano, A., Matsumoto, Y., Hasebe, N., Abe, K.,
Hayakawa, K., Ikeda, T., Iwasaki, K., Fujiwara, M., 2004. The serotonin1A agonist
8-OHDPAT reverses delta 9-tetrahydrocannabinol-induced impairment of
spatial memory and reduction of acetylcholine release in the dorsal hippocampus in rats. Neurotox. Res. 6, 153e158.
Johansson-Wallsten, C.E., Meyerson, B.J., 1994. The ontogeny of tolerance to the 5HT1A agonist 8-OH-DPAT: a study in the rat. Neuropharmacology 33, 325e330.
Kaufman, J., Delorenzo, C., Choudhury, S., Parsey, R., 2016. The 5-HT1A receptor in
major depressive disorder. Eur. Neuropsychopharmacol. 26, 397e410.
Keller, B., García-Sevilla, J.A., 2015. Regulation of hippocampal Fas receptor and
death-inducing signaling complex after kainic acid treatment in mice. Prog.
Neuropsychopharmacol. Biol. Psychiatry 63, 54e62.
Kennett, G.A., Dourish, C.T., Curzon, G., 1987. Antidepressant-like action of 5-HT1A
agonist and conventional antidepressants in an animal model of depression.
Eur. J. Pharmacol. 134, 265e274.
Kreiss, D.S., Lucki, I., 1997. Chronic administration of the 5-HT1A receptor agonist 8OHDPAT differentially desensitizes 5-HT1A autoreceptors of the dorsal and
median raphe nuclei. Synapse. 25, 107e116.
Kurisu, K., Yenari, M.A., 2018. Therapeutic hypothermia for ischemic stroke; pathophysiology and future promise. Neuropharmacology 134, 302e309.
López-Otín, C., Blasco, M.A., Partridge, L., Serrano, M., Kroemer, G., 2013. The hallmarks of aging. Cell 153, 1194e1217.
Mao, Z., Song, Z., Li, G., Lv, W., Zhao, X., Li, B., Feng, X., Cheng, Y., 2013. 8-hydroxy-2(di-n-propylamino)tetralin intervenes with neural cell apoptosis following
diffuse axonal injury. Neural Regen. Res. 8, 133e142.
Marques, F.Z., Markus, M.A., Morris, B.J., 2010. The molecular basis of longevity, and
clinical implications. Maturitas 65, 87e91.
Mattson, M.P., Magnus, T., 2006. Ageing and neuronal vulnerability. Nat. Rev. Neurosci. 7, 278e294.
McGrath, J.C., Lilley, E., 2015. Implementing guidelines on reporting research using
animals (ARRIVE etc.): new requirements for publication in BJP. Br. J. Pharmacol.
172, 3189e3193.
Mckinney, B.C., Sibille, E., 2013. The age-by-disease interaction hypothesis of latelife depression. Am. J. Geriatr. Psychiatry 21, 418e432.
Morrison, J.H., Baxter, M.G., 2012. The ageing cortical synapse: hallmarks and implications for cognitive decline. Nat. Rev. Neurosci. 13, 240e250.
Mowla, A., Mosavinasab, M., Haghshenas, H., Haghighi, B.A., 2007. Does Serotonin
augmentation have any effect on cognition and activities of daily living in
Alzheimer’s dementia? A double-blind, placebo-controlled clinical trial. J. Clin.
Psychopharmacol. 27, 484e487.
Ramos-Miguel, A., García-Sevilla, J.A., Barr, A.M., Bayer, T.A., Falkai, P., Leurgans, S.E.,
Schneider, J.A., Bennett, D.A., Honer, W.G., García-Fuster, M.J., 2017. Decreased
cortical FADD protein is associated with clinical dementia and cognitive decline
in an elderly community simple. Mol. Neurodegener. 12, 26.
Riedel, W.J., Sobczack, S., Nicolson, N., Honig, A., 2002. Stress, cortisol and memory
as markers of serotonergic vulnerability. Acta Neuropsychiatr. 14, 186e191.
Robson, L., Aj, G., Da, K., Marsden, C., 1993. Age-related behavioural, neurochemical
and radioligand binding changes in the central 5-HT system of Sprague-Dawley
rats. Psychopharmacology 113, 274e281.
Rosenzweig, E.S., Barnes, C.A., 2003. Impact of aging on hippocampal function:
plasticity, network dynamics, and cognition. Prog. Neurobiol. 69, 143e179.
Samson, R.D., Barnes, C.A., 2013. Impact of aging brain circuits on cognition. Eur. J.
Neurosci. 37, 1903e1915.
Sarubbo, F., Moranta, D., Asensio, V.J., Miralles, A., Esteban, S., 2017. Effects of
resveratrol and other polyphenols on the most common brain age-related
diseases. Curr. Med. Chem. 24, 4245e4266.
Schmitt, J.A., Wingen, M., Ramaekers, J.G., Evers, E.A., Riedel, W.J., 2006. Serotonin
and human cognitive performance. Curr. Pharm. Des. 12, 2473e2486.
Slattery, D.A., Markou, A., Cryan, J.F., 2007. Evaluation of reward processes in an
animal model of depression. Psychopharmacology 190, 555e568.
Vorhees, C.V., Williams, M.T., 2014. Assessing spatial learning and memory in rodents. ILAR J. 55, 310e332.
Walsh, R.N., Cummins, R.A., 1976. The open-field test: a critical review. Psychol. Bull.
83, 482e504.
Wolff, M., Costet, P., Gross, C., Hen, R., Segu, L., Buhot, M.C., 2004. Age-dependent
effects of serotonin-1-A receptor gene deletion in spatial learning abilities in
mice. Brain Res. Mol. Brain Res. 130, 39e48.
Wu, C., Gong, W.G., Wang, Y.J., Sun, J.J., Zhou, H., Zang, Z.J., Ren, Q.G., 2018. Escitalopram alleviates stress-induced Alzheimer’s disease-like tau pathologies and
cognitive deficits by reducing hypothalamic-pituitary-adrenal axis reactivity
and insulin/GSK-3b signal pathway activity. Neurobiol. Aging 67, 137e147.
Yoon, T., Okada, J., Jung, M.W., Kim, J.J., 2008. Prefrontal cortex and hippocampus
subserve different components of working memory in rats. Learn Mem. 15,
97e105.
Yu, H., Lewande, T., 1997. Pharmacokinetic and pharmacodynamic studies of (R)-8hydroxy-2-(di-n- propylamino)tetralin in the rat. Eur. Neuropsychopharmacol.
7, 165e172.
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