Low Plasma Immunoreactive p-Endorphin Levels in Autism

RONIT WEIZMAN, M.D., IRIT GIL-AD, PH.D., JACOB DICK, M.D., SAM TYANO, M.D.,GEORGE A. SZEKELY, M.D. , AND ZVI LARON, M.D.

Abstract. Baseline morning basa1 plasma immunoreactive /l-endorphin (ir-/l-EP) level was assayed in 22autistic, 22 schizophrenic, and 22 control subjects. Eight autistics and eight schizophrenics were drug-free whilethe remaining patients were maintained on neurolept ic treatment. The mean values of ir-/l-EP were significantlylower in unmedicated and medicated aut istics compared to age- and sex-matched schizophrenics and healthycontrols . Unaltered ir-/l-EP levels were observed in untreated schizophrenics, while neurolept ic treatment induceda significant elevation of the opioid levels. A lower ir-/l-EP elevation was achieved by neuroleptics in autisticswhen compared to treated schizophrenics. J. Am . Acad. Child Adolesc. Psychiatry, 1988, 27, 4:430-433. KeyWords: autism, schizophrenia, immunoreactive-d-endorphin, neuroleptics,

Autism is a behavioral syndrome characterized by pervasivedisturbances of interpersonal relationships, impairment incommunication, and bizarre responses to environment. Onsetis typically before 30 months of age. No single etiologicalfactor is known to be responsible for this disorder. Kalat(1978) pointed to the similarity between the behavioral attri­butes of autism and those of opiate addiction. Autistic chil­dren are socially withdrawn, sometimes insensitive to pain ,alternatively hyperactive or hypoactive, ignore sensory stim­uli, display repetitive and stereotyped behavior, manifestunexplained and inconsolable crying, and are prone to epilep­tic seizures. Panksepp (1979) demonstrated that morphinetreatment induced, in young animals, developmental andbehavioral disturbances corresponding to those observed inautistic children. Based on the analogy between opiate addic­tion and autism and the similarity between opiate-inducedpsychosocial distortion in animals and clinical manifestationsof autism, the endorphin hypothesis was postulated (Kalat,1978; Panksepp, 1979; Sandyk and Gillman, 1986).

Considering the possible involvement of the endorphinergicsystem in autism, the authors investigated in a previous studythe basal blood levels of humoral (H) endorphin in autisticpatients (Weizman et al., 1984). H-endorphin is an opiate­like endogenous ligand found in the brain, blood, and CSF ofboth humans and rats (Same et al., 1978, 1980). In contrastto the expected excess endorphinergic activity, low H-endor­phin levels were observed in autistic subjects in comparisonto those of normal controls. Decreased H-endorphin levels donot indicate the same alteration in other endorphins, sinceendorphins derive from separate precursors, are distributed in

Accepted February J. 1988.Dr. Weizman is with the Pediatric Department. Hasharon Hospital.

Petah Tiqva. Dr. Gil-Ad and Dr. Laron are with the Institute ofPediatric and Adolescent Endocrinology. Beilinson Medical Center.Dr. Dick is with the Pediatric Department. Kaplan Hospital. Rehovot.Dr. Tyano is with Geha Psychiatric Hospital. Beilinson MedicalCenter. Dr. Szekely is with Ness Ziona Governmental Hospital.Sackler Faculty ofMedicine. Tel Aviv University. Israel.

Reprint requests to Dr. Weizman. Guest Researcher. c/o NationalInstitute ofMental Health. Clinical Neuroscience Branch. NIH Bldg10. Rm 4N212. 9000 Rockville Pike. Bethesda. MD 20892.

0890-8567/88/2704-0430$02.00/0© 1988 by the American Acad­emy of Child and Adolescent Psychiatry.

discrete areas in the CNS, and act independently (lmura etal., 1985).

The observation of a physiological decline in ir-fj-EP levelsin rat brain during maturation led Panksepp (1979) to proposethe possibility that autism may reflect a failure of the striatalir-fj-EP system to exhibit this maturational decrement.

The present study was undertaken to investigate possiblealterations in blood ir-fj-EP levels in untreated and treatedautistic patients in comparison to schizophrenics and normalhealthy controls. Schizophrenics were included as a compar­ison group because of their severe psychopathology and be­cause in both disorders long-term therapy with neurolepticsis frequent. Autism is thought to be a distinct disorder withno relationship to schizophrenia, yet in the past it has beenconsidered as the earliest manifestation of childhood schizo­phrenia (Bender, 1959; Kanner, 1949), and recently, cases ofautistic children who became schizophrenics were described(Petty et al., 1984).

Method

Subjects

Sixty-six subjects aged 9 to 27 years participated in thisstudy. The study population included three groups, comprisedof 22 subjects each: autistic patients, schizophrenics, andhealthy controls. An subjects were physically healthy, non­obese, nonaddicted, and without diagnosable neurologicaldisease. Diagnoses ofautistic and schizophrenic disorder wereestablished according to DSM-III criteria. All patients werehospitalized in psychiatric units. The study was approved bythe Hospital Ethical Committee.

Autistic patients. Fifteen male and seven female autisticpatients, aged 9 to 27 years (mean ± S.D.: 16.3 ± 5.7 years),participated in the study . Eight of the 22 patients (five maleand three female), aged 9 to 15 years (11.3 ± 2.3), were drug­free for at least 6 months. The remaining 14 patients (10 maleand 4 female), aged II to 27 years (19.4 ± 4.7), were main­tained on neuroleptics for at least 6 months (eight patientswith haloperidol 15 to 30 rug/day, three with trifluoperazine15 to 30 mg/day, and three with thioridazine 150 to 300 mg/day).

430

LOW PLASMA IR-{j-EP LEVELS IN AUTISM 431

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pmol/L, p < 0.01) and the medicated autistics (5.2 ± 0.3pmol/L, p < 0.0 I) (Fig. 2).

FIG. 2. Plasma ir-fJ-EP levels in medicated autistic (N = 14) andschizophrenic (N = 14)patients vs. age- and sex-matched controls (N= 14). ••p < 0.01 vs. autistics and controls.

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FIG. I. Plasma ir-fJ-EP levels in untreated autistic (N = 8) andschizophrenic (N = 8) patients vs. age- and sex-matched controls (N= 8). •p < 0.05 vs. schizophrenics and controls.

Medicated versus Unmedicated Patients

The controls matched with the unmedicated patients(younger subjects) did not differ in their ir-fJ-EP levels fromthe controls matched with medicated patients (older subjects),despite their age difference. Ir-fJ-EP values were higher (70%)in the neuroleptic-treated schizophrenics in comparison to

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Results

Unmedicated Patients

Ir-fJ-EP levels of the untreated autistics (4.1 ± 0.3 pmol/L)were significantly lower compared to age- and sex-matchedhealthy controls (8.2 ± 1.1 pmol/L; p < 0.005) and untreatedschizophrenics (8.0 ± 1.1 pmol/L, p < 0.005). No differencewas observed between drug-free schizophrenics and matchedhealthy controls (Fig. I).

Medicated Patients

Ir-fJ-EP levels did not differ significantly in medicated au­tistics (5.3 ± 0.5 pmol/L) in comparison with their age- andsex-matched normal controls (7.1 ± 0.8 pmol/L). Medicatedschizophrenics had significantly higher ir-fJ-EP levels (13.6 ±2.0 pmol/L) than their matched normal controls (7.1 ± 0.8

Ir-fJ-EP Determination

Plasma ir-fJ-EP levels were determined using materials anda protocol derived from Immuno Nuclear Corp (Stillwater,MN, USA). IrfJ-EP was extracted from plasma using a seph­arose antibody (Krantz and Brown, 1985). Cross-reactivity ofthe fJ-EP antibody with human fJ-lipoprotein (fJ-LPH) was50% and 0.1 % with other relevant peptides (a-endorphin,dynorphin, leucine, enkephalin, methionine enkephalin,ACTH '- w, ACTH '-

24, a-MSH, fJ-MSH). Elution was per­

formed using 0.025 N HCI and elutes were tested for endor­phin using a standard double antibody RIA. The sensitivityof the assay was 2.5 pmol/L, and all samples were run in thesame assay. Intra-assay coefficient ofduplicate assays was 8%.

Statistical Evaluation

Statistical analysis of the data was performed using the one­way ANOVA. All results are expressed by mean ± S.E.M. Ir­fJ-EP levels were not disclosed to the clinicians, and diagnoseswere not available to the laboratory team until the end of thestudy.

Schizophrenic patients. Fifteen male and seven female pa­tients with an age distribution similar to the autistics (17.3 ±5.1) and suffering from schizophrenic disorder comprised thisgroup. Eight schizophrenics (five male and three female), agedII to 15 years (12.6 ± 1.2), were drug-free for at least 6months, while 14 patients (10 male and four female), aged 12to 27 years (20.1 ± 4.2), were maintained on chronic neuro­leptic treatment for at least 6 months (eight patients withhaloperidol 20 to 30 mg/day, three with trifluoperazine 15 to30 mg/day, and three with thioridazine 150 to 300 mg/day),

Normal controls. Twenty-two healthy volunteers (15 maleand seven female), aged 9 to 25 years (16.5 ± 5.4), wereincluded as controls. Eight of the subjects were matched tothe untreated autistics (11.6 ± 2.1 years) and 14 were matchedto the treated autistics (19.6 ± 3.9 years) by age and sex. Allthe control subjects were drug-free.

Procedure

Blood samples were drawn between 8:00 and 9:00 A.M. afteran overnight fast. The blood was withdrawn into standardplastic EDTA tubes and plasma was separated promptly bycentrifugation and frozen at -20· C until assayed.

432 WEIZMAN ET AL.

the unmedicated schizophrenics, although the difference didnot reach a significant level when all the groups (unmedicatedand medicated autistics and schizophrenics) were comparedwith the pooled controls (7.5 ± 0.6 pmol/L) (Fig. 3). A lessprominent difference (29%) was observed between treatedand untreated autistics. Medicated schizophrenics demon­strated significantly higher ir-fj-EP plasma levels than pooledcontrols (p < 0.01) as well as from treated and untreatedautistics (p < 0.01) (Fig. 3).

Discussion

The major finding in this study is that basal plasma ir-fJ­EP levels are reduced in untreated autistic patients whencompared to untreated schizophrenics and age- and sex­matched controls. Schizophrenics maintained on chronic neu­roleptic treatment exhibited significantly elevated plasma ir­fj-EP levels in comparison to medicated autistics and matchedcontrols. Antipsychotic agents induced a relatively slight ele­vation in ir-fj-EP in autistic patients, which was sufficient toreach the normal (but not above) levels ofthe healthy controls.

The decreased morning basal plasma ir-fj-EP concentrationin unmedicated autistics is similar to the earlier observationof reduced H-endorphin blood levels in autism (Weizman etaI., 1984). The present finding does not necessarily contradictthe opiate-excess hypothesis of autism (Deutsch, 1986; Kalat,1978; Panksepp, 1979), although it is not clear whether thedecreased blood opioid levels found reflect a parallel reductionin the activity ofthe central endorphinergic system. Peripheralopioid levels may be reciprocally related to cerebral opioid

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FIG. 3. Plasmair-tJ-EP levels in medicated and unmedicated autisticsand schizophrenics in comparison to total pooled controlsubjects (N= 22). • •p < 0.01 vs. controls as well as vs. unmedicated andmedicated autistics (but not vs. unmedicated schizophrenics).

activity. The reduced levels ofcirculating opioid may indicatehigh central opioid activity in autism (Shaley and Panksepp,1987). This possibility is supported by the finding thatsuppression of central opioid activity by intracerebral injec­tion ofnaloxone increases the circulating levels offj-EP (Levinet aI., 1984). The endorphinergic system is heterogenous withvarious independent mediators, and the decrement in thosetwo endorphins does not imply that the same mechanism isresponsible for the low levels of the two endogenous opiates.

The reduced ir-fj-EP plasma levels observed in this studyare not consistent with the reported elevated CSF ir-fJ-EP inautism and a correlation between the beneficial effect offenfluramine treatment and suppression ofCSF ir-fj-EP levels(Ross et aI., 1985). One report has shown a lack ofcorrelationbetween CSF and blood levels of fj-EP in normal subjects(Genazzani et aI., 1982). Thus, it might be that plasma ir-fJ­EP levels reflect the pituitary secretion rather than hypotha­lamic endorphinergic activity. Another study concerning CSFopioid activity in autistic children demonstrated higher meanendorphin fraction II levels in comparison to normal children(Gillberg et aI., 1985). There was a trend toward a correlationbetween high fraction II levels and decreased pain, sensibility,and self-destructiveness. The major opioid peptide in fractionII of human CSF seems to be methionine enkephalin-lys, apeptide deriving from proenkephalin A (GiUberget aI., 1985);hence this opioid cannot indicate the CSF content of fJ-EP.

Low ir-fj-EP plasma levels were observed in the presentstudy in medicated autistic patients, despite the neuroleptictreatment, which is believed to elevate ir-fj-EP plasma levels(Emrich et aI., 1980; Naber et al., 1984; Murburg et aI., 1986).The moderate response of ir-fj-EP secretion to chronic neu­roleptic administration in autistics in contrast to prominentelevation in schizophrenics might support the possibility ofhypoactivity of the endorphinergic system in autism. The lackof involvement of fJ-EP in schizophrenia is in accordancewith the reports of normal ir-fJ-EP in CSF and plasma ofschizophrenics (Emrich et aI., 1979; Naber et aI., 1981; Rosset aI., 1979) but disagrees with the finding of elevated levelsof CSF ir-fJ-EP in acute schizophrenics and low levels inchronic schizophrenics (Domschke et aI., 1979). The normalbasal morning ir-fj-EP plasma levels observed in drug-freeschizophrenics is consistent also with a recent study thatdemonstrated wide fluctuations of plasma ir-fJ-EP levelsthroughout the day but with no significant difference incomparison with normal controls (Gil-Ad et aI., 1986). How­ever, the current results contradict a report ofelevated plasmair-fj-EP levels in chronic schizophrenics (Brambilla et al.,1984). An increase in plasma ir-fJ-EP levels induced by neu­roleptics was reported in rats (Hollt, 1981) and humans(Emrich et aI., 1980; Murburg et aI., 1986; Naber et aI., 1984).

The decreasing ir-fJ-EP plasma levels both in drug-free andtreated autistics differentiate autism from schizophrenia. It isnot clear whether the hypoactivity offj-EP release mechanism,at the pituitary level, is involved in the pathophysiology ofautism. The reduced ir-fJ-EP levels in autistic patients mightaccount for some of the clinical manifestations ofautism suchas hypersensitivity to sensory stimuli, mood lability, panicreactions to minor changes, unexplained crying, and restless­ness.

LOW PLASMA IR-{3-EP LEVELS IN AUTISM 433

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