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Human antiidiotypic antibody against opiate receptors.

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Human Antiidiotypic Antibody
Against Opiate Receptors
Benjamin F. Roy, MD," Wayne D. Bowen, PhD,SJames S. Frazier, MD,? John W. Rose, MD,S
Henry F. McFarland, MD,II Dale E. McFarlin, MD,'I Dennis L. Murphy, MD,? and John M. Morihisa, MD'
Sera containing antibodies to p-endorphin from 2 patients with major depressive disorder were shown to have antiidiotypic antibodies that specifically inhibited reactivity between anti-P-endorphin I g G and p-endorphin. Autologous
and homologous antiidiotypic anti-anti-P-endorphin IgG antibodies were isolated by affinity chromatography. The
purified antiidiotypic antibody did not bind P-endorphin but competed with {'25L]P-endorphin for rat brain opiate
receptors. Normal IgG that was similarly treated had negligible competitive effects. The antibody bound to the
membrane preparation; such binding was inhibited by opiate receptor ligands. Binding of the antiidiotype to a 60,000dalton protein from rat brain was detected by Western immunoblot analysis. This protein corresponds in molecular
weight to proteins proposed to be components of opiate receptors. These findings imply that immune reactivity to
neuropeptides could contribute to psychiatric impairment.
Roy BF, Bowen WD, Frazier JS, Rose JW,
McFarland HF, McFarlin DE, Murphy DL, Morihisa JM.
Human antiidiotypic antibody against opiate receptors. Ann Neurol 1988;24:57-63
Antigen stimulates the production of antibodies. A
structural diversity of the variable region of these immunoglobulins, the idiotype, can be antigenic and
stimulate the production of antiidiotypic antibodies.
The immune network theory postulates that some
antiidiotypic antibodies can mimic portions of antigen
and potentially compete with antigen for binding to
the idiotypic antibody. Of particular relevance to the
neuroendocrine system is the concept that antiidiotypes directed at antihormone antibodies could have
the potential to bind to hormone receptors [17.
Antiidiotypes that mimic physiological characteristics of hormones or neurotransmitters have been demonstrated for the P-adrenergic receptor in asthma and
allergic rhinitis [2-4 J, the insulin receptor in diabetes
mellitus E5-71, the acetylcholine receptor in myasthenia gravis 18-10], and the thyrotropin receptor in
hyperthyroidism C11, 121. In some disease states,
idiotypic and antiidiotypic antibodies occur cyclically
[13, 141 and can potentidly reverse or arrest the progression of immunopathological abnormalities induced
by the idiotype 1151.
Recently, human autoantibody to the opioid peptide
P-endorphin has been identified by enzyme-linked immunosorbent assay (ELISA) and isolated by affinity
From the "Department of Psychiatry, Georgetown University
School of Medicine, and the Veterans Administration Medical Center, WahiWon, DC; the thborarory of Clinical Science, National
Institute of Mental Health, Bethesda, MD; the $Section of Biochemistry, Division of Biology and Medicine, Brown University,
Providence, RI; the BNeurovirology Research Laboratory, Veterans
Administration Medical Center, Salt Lake City, UT; and the
"NeuroimmunologyBranch, National Institute of Neurological and
Communicative Disorders and Stroke, Bethesda, MD.
chromatography from a subgroup of patients with major depressive disorder and somatic symptoms 1161.
This observation suggested that these patients' symptoms might reflect alterations in the activity of antibodies to opioid peptide. The mechanisms responsible
for the production of anti-@-endorphin antibody are
unknown, but might result from aberrant regulation of
idiotype-antiidiotype interactions. In this study the
possible occurrence of an antiidiotypic antibody to
anti-P-endorphin IgG has been examined.
Materials and Methods
Afinity Chromatography
IgG was isolated by affinity chromatography on protein A
(Staphylococcusaureus) from the serum of 7 normal volunteers
and 2 patients previously found to have significant amounts
of anti-@endorphin IgG and who met the criteria outlined in
the third edition of the Diagnostic and Statistical Manual of
Mental Disorders (DSM 111) for major depressive disorder.
Fresh sera from the patients with depression originally
studied [l6} were obtained; somewhat lower anti-p-endorphin antibody levels were found 3 years later. Anti+endorphin IgG (idiotype)was isolated from the IgG of the 2
patients by immunoadsorption to P-endorphin coupled to
Sepharose 4B as previously described E 163. Anti-P-endorphin antibody immunoprecipitated ['251]p-endorphinbut did
Received Aug 14, 1987, and in revised form Jan 27, 1988. Accepted
for publication Jan 3 1, 1988.
D~ R ~ chief,
~ , ~mmunologySection,
Address correspondence
School of
Department of Psychiatry, Georgetown
DC 2ooo7,
Medicine, 3800 Reservoir Road, NW,
Copyright 0 1988 by the American Neurological Association 57
not immunoprecipitate ['*'I)morphine (Roche Diagnostic
Systems, Nutley, NJ).
To determine if anti-anti-@-endorphin IgG antibody (antiidiotype) was present, human anti-@-endorphinIgG (10 mg,
idiotype) purified from Patient 1 was coupled to cyanogen
bromide-activated Sepharose 4 B (0.5 mg IgG/ml Sepharose
4B). Effluents from Patients 1 and 2 from which anti-pendorphin antibodies had been removed were immunoadsorbed to the human anti-P-endorphin IgG. After extensive
washing the putative antiidiotype was eluted. Normal IgG
was similarly treated.
F(ab')2 fragments were generated from the purified IgG of
Patient 1 as previously described [lb]. The purity of the
F(ab')z fragments was determined by double agar diffusion
(Ouchterlony) and the concentration adjusted to 5 p d 1 0 0
pl. The F(ab')z fragments were attached to an ELISA plate
and used in assay binding between whole IgG, idiotype, and
antiidiotype. After incubation for 2 hours at 37"C, binding to
F(ab')* was detected using a 1:200 dilution of goat antihuman IgG (heavy-chain specific) alkaline phosphatase conjugate.
Measles Antibody Assay
The effects of the antiidiotype on the activity of an unrelated
antibody were tested in a competitive ELISA using a human
antimeasles serum. Vero cell (African green monkey kidney
cells, American Type Culture Collection, Rockville, MD)
monolayers were infected with Edmonston strain measles
virus as previously described t17). Lysates of the infected
Vero cells were adsorbed to Immulon I ELISA plates (Dynatech, Alexandria, VA) overnight at 4°C. The concentration of antiidiotype and idiotype was 1 pg/lOO pl.
Opiate Receptor Assay
The crude mitochondrial (PL)membrane fraction of rat brain
was used for opiate receptor studies. Male Sprague-Dawley
rats (Taconic Farms, Germantown, NY) weighing 180 to
250 g were decapitated. Whole brains minus cerebellum
were placed in cold 10 mMTris-HCI, p H 7.4, containing 320
mM sucrose (Trdsucrose buffer) and homogenized in a glass
homogenizer by 10 strokes of a motor-driven Teflon pestle
in a volume of 10 mug tissue. The homogenate was centrifuged at 1,000 g for 10 minutes and the supernatants were
saved. The pellets were suspended in 2 mug ice-cold Tris/
sucrose and centrifuged again at 1,000 g for 10 minutes. The
combined supernatants were centrifuged at 3 1,000 g for 15
minutes. The pellets were resuspended in 3 mug 10 mM
Tris-HC1, p H 7.4, and the suspension was allowed to incubate at room temperature for 15 minutes. Following centrifugation at 3 1,000 g the pellets were resuspended to a final
volume of 1.53 mug in 10 mM Tris-HC1, p H 7.4. Aliquots
were stored at - 80°C until use. Protein was determined by
the method of Lowry and associates E18).
Human {'*51)@-endorphin binding was assayed using a
modification of the method described by Howard and associates [19]. To assess inhibition, purified IgG samples from
7 normal volunteers, or purified anti-@-endorphin,or antiidiotypic antibodies from 2 patients with major depressive
disorder were preincubated with 130 to 150 pg of membrane protein in 50 mM K2HP04, p H 7.4, containing 100
58 Annals of Neurology
Vol 24
No 1 July 1988
pg/ml Bacitracin, 4 cLg/ml leupeptin, 2 p d m l chymostatin,
and 10 pg/rnl bestatin. Following preincubation for 60
minutes at 25"C, 5 pl of (3-['~5SI}iodotyrosy127)@-endorphin
(human) (Amersham Corporation, Arlington Heights, IL),
approximately 2,000 CUmmol. was added, producing a final
concentration of 0.5 nM in a total volume of 100 PI. Additional incubation was carried out for 60 minutes at 25°C.
Nonspecific binding was determined in the presence of 10
p~ levallorphan, which was added at the time of radioligand
addition. All incubations were carried out in polypropylene
microfuge tubes in duplicate. To terminate the assay, a 75-pl
aliquot of the incubation mixture was filtered under vacuum
through glass fiber filters (Schleicher and Schuell, Keene,
NH). Trapped membranes were washed three times with 3
ml of cold 50 mM KlHPO4, p H 7.4. Filters were soaked at
least 30 minutes at 25°C in a solution of 50 mM K2HPO4,
p H 7.4, 0.01% poly-L-lysine HBr (M, = 500,000), and
0.4% bovine serum albumin (BSA) prior to use to reduce
nonspecific binding of ['251]@-endorphin to filters.
The inhibition due to antibodies was expressed as inhibition units per microgram of I g G in the assay. Inhibition units
were derived by determining the sample counts per minute
as a percentage of the control counts per minute and dividing
by the concentration of IgG in the assay. This facilitated
comparison of antibody effects across experiments.
Binding of Antiidiotypic Antibodies
to Rut Brain Membranes
To determine if the antiidiotypic antibody bound to rat brain
membranes, an ELISA was used. Rat brain protein (0.5 rng)
was incubated for 60 minutes ai 25°C in 10 mM K2HPO4,
p H 7.4, containing 10 PM levallorphan or
M pendorphin. T h e membranes were then centrifuged at 40,000
rpm for 10 minutes and washed and resuspended in 10 mM
Tris-HC1 buffer, p H 7.4. These solutions were then allowed
to coat the plate for 2 hours at room temperature. The plates
were washed and incubated with antiidiotypic antibody from
Patient 1, followed by incubation with secondary goat antihuman IgG (heavy-chain specificj alkaline phosphatase conjugate. All steps were carried out at room temperature.
at 90
Values reflect the optical density at 405 nm (OD4o5)
minutes after subtraction of the background binding of the
goat anti-human IgG (heavy-chain specific) alkaline phosphatase conjugate to rat brain membranes (ODdo5uniformly <
Two-Dimensional Western lmmunoblot
Proteins contained in the rat brain mitochondria1 P2 membranes (100 pg) were separated by two-dimensional electrophoresis and identified by silver staining. Gels containing
the separated proteins were also used to detect reactivity
with antiidiotype fractions from Patient 1. Two-dimensional
electrophoresis was performed according to the procedures
of OFarrell[20] with minor modifications [21}. In addition,
only pH-3.5 to -10 ampholytes were employed at a hnal
ampholyte concentration of 296. Gels to be stained with
silver were immersed in a fixative solution (50% methanol,
10% acetic acid) for 16 to 20 hours and then stained according to the procedures of Merril and associates [22] with
minor modifications {231. Gels selected for electroelution of
proteins onto nitrocellulose paper were treated according to
the procedure of Towbin and associates f24J Once protein
transfer was complete, additional antibody-binding sites were
blocked by incubating blots in 10 mM Tris buffer, p H 7.4,
150 m~ NaCI, and 3% BSA. Next, antiidiotype IgG (10 pg)
in 25 ml of buffer with 35% BSA and 4% normal goat serum was incubated for 16 to 20 hours. Blots were then incubated with peroxidase-labeled goat anti-human IgG antibody
(EGrkegaard-Perry Laboratories, Gaithersburg, MD) in 3%
BSA and 4% normal goat serum for 2 hours. Finally, blots
were colorized by the addition of 4-chloro-I-naphthol and
hydrogen peroxide. Resulting spots were then localized on
the silver-stained reference gel with the aid of transparent
Antiidiotype Inhibition of ldiotype Binding
t o PEndorphin
The capacity of the idiotype and antiidiotype to react
with @-endorphinwas determined by ELISA [lb}. The
preparations of idiotypic IgG from both patients demonstrated significant reactivity for p-endorphin (Table
1). IgG from an age- and sex-matched normal individual that had been adsorbed to P-endorphin coupled to
Sepharose 4B did not show anti-P-endorphin activity
(see Table 1).
Preincubation of idiotype from both patients with
the respective autologous antiidiotype inhibited binding of the idiotype to P-endorphin in the ELISA by
72% and 67% for Patient 1 and Patient 2, respectively
(see Table 1). The antiidiotypes from each patient
significantly inhibited the binding of the other idiotype
to P-endorphin. The antiidiotype obtained from Patient 2 inhibited the binding of Patient l idiotype to pendorphin to a greater extent (81%) than the autologous antiidiotype did. This may be related to the fact
that Sepharose-coupled idiotype from Patient 1 was
used to prepare both antiidiotypes.
Experiments were performed to exclude binding of
the antiidiotype to determinants on the Fc portion of
the idiotype. These demonstrated binding of the antiidiotype to F(ab')2 fragments of idiotypic anti-pendorphin antibody (Table 2). The idiotype from Patient l bound to the autologous F(ab')z; this was
inhibited when the idiotype was preincubated with
antiidiotype. I g G from a normal volunteer that had
been immunoadsorbed to P-endorphin demonstrated
negligible binding to Patient 1 F(ab')z.
The capacity of the antiidiotype to block the binding
of the idiotype to p-endorphin was assessed in a competitive radioimmunoassay (RIA) (New England Nuclear Research Products, Boston, MA). It was initially
shown that human idiotype (300 ng, Patient 1) inhibited binding of rabbit anti-$endorphin antiserum
to ["51fB-endorphin by 68%. The antiidiotype, when
combined with the idiotype, completely neutralized
idiotype competition with the rabbit antiserum for
Table 1. Anti-@Endorphin Activity of Human IgG
Patient lb
Idiotype + autologous
Idiotype + Patient 2
Patient zd
Patient 1
Normal volunteer'
Patient 1 I g G control
0.04 1
0.02 1
"Anti-@-endorphin activity was determined by subtracting background binding to another proopiomelanocorticotropin peptide,
corticotropin (ACTH), as described elsewhere 161. Background
binding to ACTH was less than 10% of the binding to @-endorphin.
bIgG concentrations were 1 &lo0 p1, except as noted.
'One microgram per 100 p,l of idiotype was combined with 1 &I00
4 of antiidiorype for a total concentration of 2 pg/lOO p1,vortexed,
and incubated at 4°C for 60 minutes prior to the enzyme linked
immunosorbent assay.
concentrations were 100 ngiIO0 pi.
'IgG concentrations were 100 ngil00 ~ 1 Patient
1 IgG was equilibrated to 100 ng/lOO PI.
= optical density at 405 nm.
Table 2. Idzotype Binding to F(ab')z Fragments
F(ab')2 Fragmentsa (ODdos)
Patient 1
Normal volunteer
0.671 0.144
0.152 0.261
0.17 1
0.04 1
"Values reflect the means of triplicate optical density at 405 nm
(OD,,,) readings minus the background binding of the antibody
conjugate to F(ab'),.
binding to {'z51]@-endorphin.In contrast, 300 ng of
antiidiotype and a normal IgG had negligible effects
(average net cpm: control = 3,027; idiotype = 986;
antiidiotype = 10; idiotype
antiidiotype = 20; normal IgG = 3,500). This experiment demonstrated interference of idiotypic antibodies in the RIA, confirmed binding of the idiotype to @-endorphin, and
provided additional evidence that the antiidiotype does
not react with @endorphin.
Roy et al: Human Antiidiotypic Antibody
Table 3. Efect of Anti-PEndarphin ldiotype and Antiidiotype
on Antimeasles Antibody Activity"
Competing Antibody
Controlb Antiidiotype Idiotype
(OD/,os) (OD/,os)
Measles antiserum titer
1: 200
1 : 400
1 : 800
1 : 1,600
1: 3,200
1 :6,400
0.46 1
"Values represent the average of duplicate optical densiry at 405 nm
(OD,iDs)readings, at 20 minutes' reaction time.
bControl is the binding of the antimeasles serum to measles-infected
Vero cells in the absence of competing antibody.
The specificity of the antiidiotype for idiotypic antiP-endorphin antibody was evaluated in a human antimeasles assay. Neither the antiidiotype nor the
idiotype significantly inhibited the reactivity between
antimeasles IgG and Vero cells infected with Edmonston strain measles virus (Table 3 ) . Additionally,
neither antibody inhibited the immunoprecipitation of
['251]morphine by a goat antimorphine antiserum (unpublished data).
Antiidiotype Inhibition of {'251)P.Endorphin
Binding to Rat Brain
The possibility that the antiidiotype cross-reacted with
P-endorphin for opiate receptor binding was studied
with human {"51]P-endorphin as radioligand and rat
brain membranes as receptor source. In each experiment, antiidiotype reduced binding of [ "'I]Pendorphin to receptors. In experiment 1 (Table 4),
antiidiotype suspended in 5%> fetal calf serum inhibited binding by 35% (0.45 inhibition unitslpg IgG).
Idiotype (anti-@-endorphin IgG) reduced binding by
60%. This presumably occurred by direct binding of
anti-P-endorphin IgG to [ 'L51)P-endorphin,reducing
the free radioligand concentration available to receptors. Depletion of IgG sample idiotype and antiidiotype produced IgG with markedly reduced ability
to inhibit C1251)P-endorphin binding (0.09 inhibition
unitslpg IgG). As fetal calf serum itself had an inhibitory effect on binding (not shown), experiment 2 was
carried out with antibodies suspended in distilled water. In experiment 2, a greater inhibitory potency of
the antiidiotype was revealed. The inhibitory potency
of patient antibodies was compared with that of IgG
from normal volunteers. Although IgG from normal
volunteers produced slight inhibition (0.32 inhibition
unitsipg IgG), antiidiotype from Patient 1 had 12.2
times the inhibitory potency (3.90 inhibition unitsfpg
IgG) of normal IgG. The combination of antiidiotype
60 Annals of Neurology Vol 24
No 1 July 1988
with idiotype mutually neutralized the inhibitory effect
of these antibodies on the binding assay (0.51 inhibition unitslpg I&), and IgG depleted of both antiidiotype and idiotype had only 11% of the inhibitory
potency of antiidiotype (0.44 inhibition unitslpg I&).
Patient 2 demonstrated 16 times the inhibitory potency (5.18 inhibition unitsfpg IgG) of normal IgG.
The incomplete inhibition observed in these assays
may suggest that subsaturating concentrations of antiidiotype were used in these experiments.
Antiidiotype Binding t o Rat Brain Membranes
Parallel experiments were conducted to determine if
binding of the antiidiotype to rat brain membranes
occurred that would account for the inhibition of
['251]P-endorphin binding. An ELISA plate coated
with the P2 fraction of rat brain was used to assay
binding of several concentrations of antiidiotype. Antiidiotype (100 nglml) bound to rat brain membranes
(Table 5). This could be inhibited by prior incubation
of rat membranes in
M levallorphan (465%reduction),
M P-endorphin (72% reduction), or the
addition of soluble @-endorphinto the antiidiotype to
compete for opiate receptor sites (84% reduction).
Control immunoglobulins showed negligible binding
to brain membranes. Thus, the antiidiotype reacted
with a component of rat brain membranes in a way that
would explain the inhibition of { '"I}P-endorphin
binding. The detection of the binding of the antiidiotype to rat brain membranes occurred at nanogram
concentrations in the ELISA that were 50-fold less
than the concentration required for inhibition in the
opiate receptor assay. Thus, the antitdiotype bound to
membranes at physiological concentrations.
IdentiJcation by Antiidiotype of a Protein
fram Rat Brain Membranes
Western immunoblot analysis was used to determine
which proteins in the rat membrane preparation (silver
stain, Fig A) were reacting with the antiidiotype. Immunoblotting with each antiidiotype revealed reactivity to a diisomeric protein with a molecular weight of
60,000 and a PI of approximately 7.0 to 7.1 (Fig B).
These data indicate the existence of a naturally occurring human antiidiotypic anti-anti+-endorphin IgG
antibody with the ability to bind to the opiate receptor.
This conclusion is supported by three lines of evidence. First, the antiidiotype inhibited the binding of
idiotypic human anti-a-endorphin IgG to 0-endorphin
in the ELISA and to soluble P-endorphin in a competitive RIA for @-endorphin.The antiidiotypic antibody
had no effect on the binding of an unrelated goat antiserum to morphine or a measles antiserum to viral
antigen in measles-infected Vero cells. Second, the
Table 4 . Eflect of Antiidiotype on Binding of {'2'I}pEndorphin t o Rat Brain Membranes
IgG (1*d100 111)
Inhibition (%)
Experiment 1'
Patient 1
IgG depleted of idiotype
and antiidiotype
Experiment 2d
Normal IgG (N
Patient 1
= 7)
IgG depleted of idiotype
and antiidiotype
Patient 2
"Inhibition units for each sample were derived by determining the sample counts per minute as a percentage of the control counts per minute
and then dividing the percentage inhibition by the concentration of IgG.
bThe normal IgG value was the mean obtained with antibodies from 7 healthy volunteers. This value (inhibition unitdpg IgG) was set at 1.0 and
patient values were expressed as a ratio to this value.
'Idiotypic and antiidiotypic antibodies were suspended in 5% fetal calf serum (FCS). The control was 5% FCS. IgG concentrations were
equilibrated by radial irnmunodiffusion. For unknown reasons, FCS depressed control binding of [1251)P-endorphinby 15%. Specific binding in
the control was 6,558 cpm after subtraction of nonspecific binding in the presence of 10 p~ levallorphan.
dAntibodies were suspended in distilled, deionized water. The concentration of rat brain membranes was constant. The ability of patients'
antibodies to inhibit 0-endorphin binding was compared with that of I g G from normal volunteers. The specific binding of the control was 12,244
cpm. The mean specific binding 5 standard deviation for the 7 normal volunteers was 10,210 2 1,199.
Table 5 . Antiidiotype Binding to Rat Brain Membranesa
Brain plus 10 p i
Levallorphan (ODdo5)
Brain plus
P-Endorphin (ODdO5)
Patient 1
Antiidiotype + l o p 5M
Normal volunteer IgG
0.103 t 0.017
0.073 2 0.055
"Values reflect optical density at 405 nm (OD405)at 90 minutes after subtraction of the background binding of the goat anti-human I g G (heavychain specific) alkaline phosphatase conjugate to rat brain membranes (uniformly ODdO5c 0.070).
antiidiotype inhibited the binding of the antigen (pendorphin) to the opiate receptor. Third, antiidiotype bound to the rat brain membrane preparation
described. Pretreatment of rat brain with either (3endorphin or the opiate antagonist levallorphan or the
addition of soluble @-endorphinto the antiidiotype inhibited binding of the antiidiotype to rat brain. These
observations suggest that antiidiotype recognizes an
antigen that also recognizes opiate receptor ligands,
presumably opiate receptors. The activity of this human antiidiotypic anti-opiate receptor antibody parallels the activity of antiidiotypic antibodies to the opiate
receptor experimentally induced to guinea pig and rabbit antibodies to morphine [25, 261.
The mechanism by which antiidiotype inhibits the
binding of ['251)p-endorphin is not known. Antiidiotype may directly occupy the ligand binding site of
the receptor. Alternatively, the antiidiotype may bind
to a nearby site and cause inhibition by inducing a
conformational change. That antiidiotype recognizes a
brain protein was confirmed by immunoblot analysis of
rat brain membrane proteins. The antiidiotype bound
to a diisomeric protein of molecular weight 60,000.
This protein may correspond to a 65,000 molecular
Roy et al: Human Antiidiotypic Antibody
Sohbilized proteins ofthe P2 fraction o j rat brain membranes
(100 p g i =‘ere identzjed by silver staining ofu tuwdimen~ional
Ref (A).The untiidiotype (10 pgi from Patient I demonstrated
binding to a diisomerir-protein of molecdar u:right 60,000 and
p l 7.0 to 7.1 (B). ImniunoblotJ ofuntiidiotypefrom Patients I
and 2 were irtenticul to the example provided The arrows in
each punel designate the hution of the diisomericprotein. These
observations were confirmed in three experiment.r.
weight protein to which @-endorphin can be crosslinked [ 19) and that is believed to be a component of
the mu and delta opioid receptors. In addition, a mu
opiate receptor component of molecular weight
58,000 has been purified from rat brain {27), and a
delta opiate receptor component of molecular weight
58,000 has been purified from NG108-15 neuroblastomaglioma cells 1281. The multiple isomeric forms of
the protein suggest the presence of a functional component, for example, a phosphorylated variant of the
Although the observed inhibition of [‘251]P-endorphin binding to rat receptor by antiidiotype was
significant, it did not represent complete inhibition.
Several explanations are possible. The moderate inhibition might have resulted from antibody being directed at determinants that are not directly associated
62 Annals of Neurology
Vol 24
No 1 July 1988
with the ligand binding site, but that have indirect effects on ligand binding. Similar reactivity for antibodies to the acetylcholine receptor have been described 191. Alternatively, the incomplete inhibition
may relate to species specificity. For example, the avidity of the human antibody for rat opiate receptor may
be relatively low, as was observed in studies of the
binding of human thyroid-stimulating immunoglobulins to mouse thyroid tissue [12). 6-Endorphin binds
to both mu and delta receptors in rat brain [29], and
there is the possibility that moderate inhibition in this
assay could have resulted from reactivity of the antiidiotype for only one receptor subtype. Finally, there
is the possibility that the concentrations of antiidiotype
used were insufficient to saturate receptor sites in the
assays described. These last two issues could possibly
be resolved by competition studies with selective mu
and delta receptor probes and Scatchard analysis of
antibody binding to membranes.
Detection of these antibodies in patients with major
depressive disorder has prompted consideration of the
possibility that abnormal regulation of idiotypic networks for opiate peptides may contribute to the
pathogenesis of affective disorders. Idiotypic and antiidiotypic antibodies to 6-endorphin and the opiate receptor may function in a receptor-specific immune network that modulates central nervous system function.
Thus, an autoimmune mechanism that perturbs the
dynamic steady state of the immune network for a
specific brain peptide might induce selective modulation or denervation of peptidergic neurons and their
Hypotheses for opioid mechanisms in psychiatric
disorders have postulated simple excess or deficiency
of opiate peptides {30, 31). Consequently, the relationship of opioid mechanisms to psychiatric disorders
has been investigated by several approaches that have
been either nonconclusive or negative, and have
militated against the participation of opioid mechanisms in affective disorder E32, 331. One method has
been the direct measurement of opiate peptide levels
or activity (by RIA or radioreceptor assay, respectively) in serum or cerebrospinal fluid. Parenthetically,
studies using direct measurement did not exclude or
control for the interference of both idiotypic and antiidiotypic antibodies in RIA or in opiate receptor assays. Thus, the presence of anti-p-endorphin or antiidiotypic antibody might lead to erroneous conclusions
about the clinical role of the opioid system in affective
and other psychiatric disorders.
An idiotypic network of this type would have the
potential to parallel and interact with a similar neuroregulatory network within the central nervous system.
Idiotypic and antiidiotypic antibodies ordinarily react
with idiotypic and antiidiotypic receptors on T and B
cells. The identification by antiidiotype of a protein
in rat cortex that presumably cross-reacts with the
idiotype of an immunoglobulin raises additional questions as to the conservation of receptor proteins, as
weU as the physiological mechanisms operating between the immune and nervous systems. There is the
possibility that the neuroregulation of neuropeptide
and neurotransmitter receptors and neural responsiveness to stimuli are modeled on self-recognition in a
manner analogous to immune network interactions.
Idiotype cross-reactive proteins might provide a
homeostatic mechanism for modulatory interactions
between the immune and nervous systems. A disturbance in the interaction between parallel networks
might induce reciprocal compensatory changes.
We would like to thank Michelle M. Dauphin, Susan B. Hellewell,
and Dr David M. Jacobowitz for technical advice and assistance, and
Mrs Dottie Drake for preparation of the manuscript.
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Roy e t al: H u m a n Antiidiotypic Antibody
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antiidiotypic, opiate, receptors, antibody, human
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