Journal of Autism and DevelopmentalDisorders, VoL 9, No. 2, 1979

Overview of Selected Basic Research in Autism L. R. Piggott Lafayette Clinic, Detroit

Basic research in autism is reviewed. There is mounting indication, but as yet inconclusive evidence, o f unique physiologic disturbances etiologically related to autism. Additionally there is indication that some o f the physiologic disturbances f o u n d in autistic children are also present in children with other developmental disorders. Children called autistic probably represent a complex o f clinically similar manifestations in a variety o f different subgroups o f children, each subgroup representing a basically different physiologic disturbance. However, the possibility remains that there is only one basic disturbance that in varying degrees affects many body systems and thus manifests in a variety o f overlapping syndromes. Objective markers are needed so as to allow the demarcation o f subgroups o f autistic children f o r further study. Possible markers may be decreased duration o f postrotatory nystagmus, auditory evoked response deviations, lymphocytic hyporesponsivity, increased blood platelet serotonin efflux, and/or the presence of urinary D M T or bufotenin.

INTRODUCTION

Autism became a popular term in child psychiatry in the years following Kanner's report of children that he referred to as having Early Infantile Autism. Initially Early Infantile Autism came to be considered an entity probably of psychological origin related to early rearing practices. The approaches used were mainly psychotherapeutic with the idea that if the autistic shell could be breached one would find a potentially normal intact child. As time passed there has been considerable disillusionment with the psychotherapy approach as well as increasing recognition that the term autism has been applied to a multiplicity of etiologically separate or interrelated conditions. Recently there has also been a dawning recognition that many children designated as autistic may well overlap with children who 199 0 1 6 2 - 3 2 5 7 / 7 9 / 0 6 0 0 - 0 1 9 9 5 0 3 , 0 0 / 0 9 1979 Plenum Publishing C o r p o r a t i o n

200

Piggott

have other forms of developmental disabilities. This overlap in some cases may include similar etiologies and in other cases may be only an overlap in symptoms. This is still unclear and will require much more extensive observation and research in the field of developmental disabilities, which can now be seen as including autistic, schizophrenic, learning-disabledl and language-disturbed children as well as those children having only mild developmental deviations. This paper reviews many of the basic studies that indicate that there may be multiple underlying organic disturbances and thus multiple organic etiologies to the syndrome known as autism. It also touches on the idea that a number of the physiologic disturbances found in autistic children are not unique to them but are also found in children with other developmental disturbances.

PHYSICAL DEVIATION STUDIES A number of investigators have reported finding physical irregularities related to childhood psychosis: placental irregularities in infants born to schizophrenic mothers (Mednick & Schulsinger, 1968), increased number of minor congenital abnormalities in schizophrenic children (Goldfarb, 1967; Quinn & Rapoport, 1974). Steg and Rapoport (1975) reported finding significantly more minor physical anomalies as scored by Waldrop's system (Waldrop, Pedersen, & Bell, 1968) in a group of severely emotionally disturbed and a group of learning-disabled children than in a group of children in a pediatric hospital, or in a group of neurotic children. They noted that the physical features in which the minor physical anomalies are manifest form during the first 3 months of gestation and that their formation may parallel the early development of the central nervous system. From this they speculated that minor anomalies may also be laid down simultaneously in the central nervous system (CNS) and that there may be basic physiologic relationships between learning disabilities and severe emotional disturbance. Goldfarb (1974) reported a 3-year longitudinal study of 40 schizophrenic children who had been admitted to Ittleson Center. Out of 40 measures he found 5 that remained unimproved (unchanged) over the 3-year span. They were (1) muscle tone, (2) activity level, (3) auditory startle response, (4) delayed auditory feedback, and (5) double simultaneous stimulation (homologous with eyes open). He suggested that the deviations in the first four areas may be related to the level of integrity of the schizophrenic child's nervous system. He further suggested the possibility that these four measures may relate to self-monitoring and/or orienting feedback systems impairments. Hauser, Delong, and Rosman (1975) found ventriculographic evidence of temporal horn enlargement, particularly in the region where the hippocampus usually bulges into the ventricle, in 15

Overview of Research in Autism

201

autistic children. This was bilateral in 5 cases and unilateral in the left ventricular temporal horn in 10 cases. They noted that the hippocampus is very sensitive to hypoxia and that in adults hippocampal damage is known to result in retrograde amnesia, a profound inability to store new knowledge, and/or emptiness of affect, one can speculate that some autistic children showing similar disabilities may have sustained hippocampal damage secondary to birth hypoxia. Further, one can reflect on possible relationships with those learning-disabled children who show definite memory disturbances. Darby (1976) reviewed reported cases of children who had been diagnosed as psychotic or autistic, had died, and had been autopsied. In all he found 33 cases. Neuropathology had been found in 27 of the 33 cases. Nine had specific diagnostic findings--7 had cerebral lipidosis, 2 had tuberous sclerosis--and 18 had nonspecific findings including brain atrophies and degenerative changes. These studies are beginning to provide a firm foundation to the belief that there is some as yet poorly understood physiologic deviation or deviations underlying the symptoms we have come to associate with the diagnosis of autism. It also appears that at least some of the physiologic deviations may have been laid down in a deviant manner during gestation. They may involve the temporal, particularly the hippocampal, area as well as other brain areas, especially those involved in feedback functions. At this point one can only speculate as to the relationship between the physiologic deviations underlying the autistic child's disturbances and the physiologic deviations that may underlie the deficits seen in children with various learning disabilities.

NEUROPHYSIOLOGIC STUDIES

Electroencephalograph&Studies EEG studies have been carried out on psychotic children by a number of investigators. White, DeMyer, and DeMyer (1964) reported finding EEG abnormality in 51% of the 149 emotionally disturbed children. The psychotic children and the nonpsychotic severely acting-out behaviorally disturbed children had significantly more EEG abnormality than the neurotic children or than a control group of 13 normal children. H u t t , Hutt, Lee, and Ounsted (1965) on recordings of EEGs in 10 autistic children found primarily low voltage irregular activity with no established rhythms. These were similar in appearance to the desynchronized EEG said to be associated with high arousal states in adults. As the environmental situation around the autistic child became more complex, their EEGs showed increasing desynchronization. Their stereotypic behavior also increased. Hutt

Piggott

202

et al. theorized that these autistic children were in a chronic high arousal state and that thee stereotypies may act as a safety mechanism to prevent further sensory input. Creak and Pampiglione (1969) found no uniformity in type, distribution, or severity of EEG abnormality in 35 children who met the British Working Party criteria for the schizophrenic syndrome. Twentynine of the 35 did have EEG abnormality. Gubbay, Lobascher, and Kingerlee (1970) found that 65% of 23 psychotic children showed EEG abnormalities but there were no characteristic EEG findings. Ritvo, Ornitz, Walter, and Hanley (1970) found that about 35% of 184 hospitalized emotionally disturbed children had abnormal EEG recordings irrespective of clinical diagnosis (psychotic or nonpsychotic) if there were specific CNS findings or a history of conditions known to be associated with organic brain dysfunction. Small (1975), using quantitative EEG analysis did find high sessionto-session positive correlation of mean EEG amplitude measures, and prominent periodic elements in autistic but not in normal children. The normals also differed from the autistics in that they showed higher mean voltage over the left hemisphere as compared to the right. This may indicate a failure of lateralization, something that is also clinically seen in a variety of children with development deviations. As yet there is no EEG finding unique to autism, although improving technology may in the future give us the tools with which to find disturbances uniquely related to some groups of autistic children. Small's studies might well be expanded to children with a variety of disturbances to see if her findings relate only to autism or to a wider range of disturbance.

Sleep Studies Sleep studies have shown some differences between autistic and normal children. In REM sleep autistic children had significantly more spindlelike (10.5 to 15 c/sec) activity and significantly fewer single eye movements and eye movement bursts than normal children (Ornitz, Ritvo, Brown, La Franchi, Parmelee, & Walter, 1969). Ornitz et al. felt that this might indicate a deficiency in sleep stage differentiation in the autistic children. Tanguay, Ornitz, Forsythe, and Ritvo (1976) compared a group of autistic children (ages 36-62 months) to an age-matched group of normals and to a group of normal children below 18 months of age. They found no significant difference,between the autistics and the normals who were below 18 months of age. The autistic children did differ significantly from their age-matched controls on mean eye movement burst duration and eye movements within bursts to eye movements outside of bursts ratio. This they felt indicated brain mechanism immaturity in the autistics. They referred t o works of Magherini, Pompeiano, and Thoden (1971) and Jeannerod,

Overview of Research in Autism

203

Mouret, and Jouvet (1965) to show that abnormalities in eye movement during REM sleep may reflect disturbance at a number of sites and levels in the CNS. These studies are beginning to suggest maturational deviations in autistic as compared to normal children. E v o k e d Potential Studies

Maturational deviation in auditory evoked response (AER) has also been indicated in studies by Ornitz, Ritvo, Panman, Lee, Carr, and Walter (1968) and by Tanguay (1976). Ornitz et al. reported that autistic children below the age of 5 years did not show a decrease in amplitude of AER during REM sleep as did a comparably age-matched group of normal children. The differences were particularly evident during eye movement burst portions as compared to ocular quiescent portions of REM sleep. They noted that in normals the inhibition of AER during the eye movement burst portions of REM sleep has been shown to be mediated by the vestibular nuclei. They related their REM findings to clinical observations suggesting possible vestibular dysfunction in autistic children. Tanguay found significantly larger auditory evoked responses over the right hemisphere as compared to the left hemisphere in normal children but no consistent hemispheric differences for AER in autistic children. This he felt may indicate a definite disturbance in the development of functional lateralization in the autistic child. Small (1971) reported finding less complex AERs and visually evoked responses (VERs) that were of low amplitude, had shorter peak latencies, and were more stable in autistics than in normal controls (N = 5 sexage matched autistic-normal pairs). When auditory and visual stimuli were presented together, the VER of the autistic children was more variable than the VER of the normal children. In 1975 she reported on an extension of this study. The only finding that remained significant was the decrease in negative amplitude of the midline recorded VER in autistic as compared to normal children. Lelord, Laffont, Jusseaume, and Stephant (1973) coupled sound and light inputs to measure the coupling effects on the AER in autistic children. First, they gave the auditory stimulus alone and measured the AER, then they gave a strong light flash stimulus 700 msec after the auditory stimulus and measured the AER. Next, they again gave the auditory stimulus alone and measured the AER. They found that the AER was stable and regular in normals but irregular and variable in autistics, that the amplitude of the AER was smaller in the autistics than in the normals, and that the normals but not the autistics showed an increase in amplitude and a diminution in variability of the AER when the auditory stimulus was coupled with the visual stimulus. They felt these findings may indicate a defect in integration between the visual and auditory pathways in autistic

Piggott

204

children. In 1975, Saletu, Saletu, Marasa, Mednick, and Schulsinger were able to differentiate children born to schizophrenic mothers (high risk) from children born to nonschizophrenic mothers correctly in 75-82% of cases using a stepwise discriminant function or a canonical correlational analysis of their auditory evoked responses. They noted that the differences found between the high risk and normal children were similar to those they had observed between psychotic and normal children and between chronic schizophrenic and normal adults. Thus they believed they might have a means to detect reasonably accurately a neurophysiologic aberration in future schizophrenics long before the onset of clinical manifestations of schizophrenia. This merits further exploration. Student and Sohmer (1978) found longer auditory nerve transmission latency and longer brainstem transmission time for the auditory evoked response in a group of 10 autistic children as compared to 12 normal children. Another 5 autistic children showed previously unsuspected definite peripheral cochlear hearing loss when their AERs were measured. It appears that definitely different REM sleep eye movement and auditory evoked response patterns are being found in children who meet various criteria for the diagnosis of autism as compared to patterns seen in age-matched normal control children. Both the auditory evoke d response and REM sleep eye movement patterns are very similar to those seen in much younger normal children. Thus one can speculate on the role that physiologic maturation plays in the development of the clinical picture we call autism. Student and Sohmer's findings are especially exciting as they more directly measure possible disturbance in function of specific nerve and brain areas. This and the ability of Saletu et al. to accurately differentiate the high risk children by AER patterns give hope that evoked response patterns may in time become diagnostic of a specific subgroup of autistic children.

Autonomic Studies

Disturbances in autonomic function have been found in a number of children given autistic or psychotic diagnoses. Hutt, Forrest, and Richer (1975) found significantly more arrhythmia in autistic children than in normal children when they recorded their heart rates in situations varying from no external stimulation to an adult actively attempting to involve the child in a task. Bernal and Miller (1970) found lower mean galvanic skin resistance responses during photic or tonal habituation trials for autistic as compared to normal children. Piggott, Ax, Bamford, and Fetzner (1973) found that a group of psychotic children did not sustain the acceleratory phase of the cardiac cycle following inspiration as long as did the controls,

Overview of Research in Autism

205

that the lag intervals between respiratory changes and related cardiac rate changes were more consistent for the controls, and that the direct relationship between rate change was lost at maximum respiratory depths for the psychotics but not for the controls. These studies indicate physiologic differences related to or reflected through autonomic control mechanisms in the psychotic as compared to the normal child.

Vestibular Studies

Goldfarb (1974) has postulated faulty self-monitoring or feedback systems to account for his clinical findings. DeMyer, Schwier, Bryson, Solow, and Roeske (1971) also suggested faulty feedback at either a neurogenic or a cellular level to account for their findings of significantly greater plasma-free fatty acid variability in a group of psychotic children as compared to nonpsychotic emotionally disturbed and normal children. One can question to what degree faulty self-monitoring accounts for the disturbances we see in autism and to what degree and manner it is also present in children with other developmental deviations. The vestibular system is believed to be intimately involved with self-monitoring functions. A number of studies have shown a decrease in duration of nystagmus after vestibular stimulation by rotation or by caloric irrigation in autistic as compared to other children (Pollack & Krieger, 1958; Colbert, Koegler, & Markham, 1959; Ritvo, Ornitz, Eviatar, Markham, Brown, & Mason, 1969). Ritvo et al. (1969) also reported that the shortened duration of postrotatory nystagmus in autistic children when tested under lighted conditions was not present when they were tested in the dark. Ornitz, Brown, Mason, and Putnam (1974), on further investigating Ritvo's finding, tested children under conditions allowing (1) light input and eye fixation, (2) light input but no eye fixation, (3) minimal light input and eye fixation, and (4) no light input and no eye fixation. They found significantly decreased duration of nystagmus and total number of nystagmic beats in the autistic as compared to normal children under conditions in which the children could fixate and had minimal light input as well as in situations in which the child could not fixate but did have light input. There were no differences when there was neither light input nor ocular fixation. Thus they demonstrated that factors related to both ocular fixation and to light input were operative in inhibiting the postrotatory nystagmus responses of autistic children. Fish (1975) reported a decreased duration of calorically induced nystagmus during the first months of life in four underactive and hypotonic infants born to schizophrenic mothers. She found that the decreased nystagmus responses were associated in these children with periods of pan-developmental retard-

206

Piggott

ation between birth and 2 years of age, with the "quiet state" in the 1st month of life, and with failure of bimanual skills in the 4th and 7th month of life. The decreased nystagmic responses were transitory, which she felt indicated CNS integrative disruption associated with development rather than with an organic vestibular lesion. Piggott, Purcell, Cummings, and Caldwell (1976) measured postrotatory nystagmus after rotation at three different rates in 25 emotionally disturbed children who were divided into 2 groups on the basis of performance on a cognitive-perceptual-motor (CPM) test battery. Those children with one or more areas of marked deficit were compared to those children with two or more areas of marked deficit. The latter group of children had a significant decrease in duration of postrotatory nystagmus after the slowest rate of rotation (30~ but not after the two faster rotation rates (60~ 90~ and a significant decrease in amplitude of postrotatory nystagmus after all rates of rotation as compared to the children with one or no areas of CPM deficit. Freeman, Frankel, and Ritvo (1976) found that autistic children, on being given a reward of different rates of rocking (vestibular stimulation) for button pushing, pushed the button more frequently as the rocking rate increased, whereas retarded nonautistic children pushed the button at the same frequency regardless of rocking rate. The overall amount of rocking each group received was the same but one can speculate as to whether the difference in rate of button pushing represents a difference in vestibular experience. At this point it appears that evidence is accumulating that indicates physiologic disturbance possibly involving the vestibular system in some children diagnosed as autistic or schizophrenic. The disturbance or disturbances, however, may not be unique to the psychotic group. The possibility that vestibular system disturbance may be intimately involved in the etiology of the clinical picture called autism is forwarded by the findings of REM sleep eye movement disturbance and auditory evoked response disturbance during REM sleep, and by postrotatory and calorically stimulated eye movement abnormalities. One can speculate on the degree to which the intactness of the vestibular system is absolutely necessary for our ability to properly perceive feedback of our own actions as well as to accurately perceive stimuli from external sources. Is this system's intactness absolutely necessary for us to be able to develop stable self- and object images? Could its dysfunction result in the perceptual inconstancy that Ornitz and Ritvo (1968) see as underlying the development of clinical autism? Could its dysfunction underlie the persistent deviations described by Goldfarb (1974) that he felt might relate to feedback systems impairments?

Overview of Research in Autism

207

BIOCHEMICAL STUDIES

Catecholamine Studies Perry (1963) found N-methylmetanephrine in the urine of 3 of 18 juvenile psychotics as compared to none of 15 mentally defective and 20 normal children. Coleman, Campbell, Freedman, Roffman, Ebstein, and Goldstein (1974) found a minor elevation of dopamine-/3-hydroxylase (DBH) serum levels in a mixed group of autistic and schizophrenic children as compared to normal controls matched for age and sex. Goldstein, Mahanand, Lee, and Coleman (1976) reported on the extension of the study of Coleman et al., which now included 78 autistic and 78 age- and sexmatched normal controls. The autistic group showed a significantly lower serum DBH activity level. Lake, Ziegler, and Murphy (1977) using the same diagnostic criteria as Coleman et al. and Goldstein et al. found decreased serum DBH but elevated norepinephrine levels in 11 autistic children and their families. The DBH levels were significantly lower in the autistic children and in the relatives of the autistic children than they were in the normal controls. Interestingly the autistic children also had significantly increased norepinephrine levels as compared to the normal controls. Since DBH is the enzyme that converts dopamine to norepinephrine, Lake et al. pointed out that one might have expected decreased rather than increased norepinephrine levels. Lake et al. noted that significantly lowered serum DBH activity levels are also found in children with Down's syndrome but their families have normal DBH activity levels. They went on to say that thus far the pattern of low DBH activity in both patient and family has been described only in autistics. This, then, is perhaps an indication of some heritable genetic deviation. Thus low serum DBH activity levels in autistics have been reported and confirmed by two investigative groups using the same diagnostic criteria. It would appear that this finding is well worth pursuing, particularly as it is believed that the catecholamine and indoleamine systems may be in dynamic balance and that disturbances in one or both systems may well be involved in childhood and/or adult schizophrenia.

Indoleamine Studies A number of studies have been carried out that focus on indoleamine, particularly serotonin metabolism, as an area of possible disturbance in childhood psychosis. Attempting to effectively study, measure, or monitor

208

Piggott

CNS serotonin function is exceedingly difficult. Serotonin does not cross the blood-brain barrier. Thus serotonin found in body fluids outside the CNS originates outside the CNS. In addition about 85-90% of the body's serotonin originates in the gastrointestinal system and only about 12% originates in the central nervous system. Therefore, when one attempts to measure the metabolic breakdown products of serotonin in various b o d y fluids, by far the preponderance of those breakdown products are from serotonin produced outside the CNS. In 1961, Schain and Freedman found elevated blood serotonin levels in 6 of 23 autistic children. All were also markedly retarded. This finding has been confirmed by other groups (Ritvo, "Yuwiler, Geller, Ornitz, Saeger, & Plotkin, 1970; Goldstein, 1976; Partington, Tu, & Wong, 1973) but it is not limited to autistic children. Campbell, Friedman, DeVito, Greenspan, and Collins (1974) found no significant differences in blood serotonin levels in psychotic and nonpsychotic grossly neurologically disturbed children. Children with lower IQs in both groups had higher blood serotonin levels than the more intelligent group members. Nonautistic retarded children also show elevated blood serotonin levels (Tu & Partington, 1972; Pare, Sandier, & Stacey, 1959). Campbell, Friedman, Green, Collins, Small, and Breuer (1975), however, have reported finding significantly increased blood serotonin levels in autistic children who were floridly psychotic as compared to autistic children who were in remission or partial remission. Narasimhachari reported on a series of studies originating in Himwich's lab (Narasimhachari & Himwich, 1975). They found 5-hydroxy N-N dimethyltryptamine (bufotenin) in the urine of 6 autistic children on from one to six occasions. None was found in the urine of 6 normal controls. Then 24-hour urine collections were done daily for 2 weeks on the 6 autistic children. Bufotenin was found in the urine of 5 of the 6 and N-N dimethyltryptamine (DMT) was found in the urine of 3 of the 6 autistic children. The urine from the parents of the 5 autistic children showing urinary bufotenin was examined. One or both parents of all 5 children showed bufotenin in their urine. Additionally 4 of 6 grandparents tested showed urinary excretion of bufotenin. Next, they blindly studied 24-hour urine samples from 23 children with a possible diagnosis of autism and from 13 young adults. Bufotenin was found in the urine of 9 of the 23 patients and 1 of the 13 controls. They concluded that determination of whether their findings indicated a genetic origin for autism and that the relatives were carriers of a metabolic defect or whether the presence of DMT and/or bufotenin in the urine is not related to a disease condition requires further investigation. Piggott, Frohman, Ward, and Gottlieb (1975) reported that the uptake of tryptophan (a serotonin precursor) by chicken red blood cells was significantly increased in the presence of serum from children meeting

Overview of Research in Autism

209

the DeMyer-Churchill criteria (DeMyer, Barton, DeMyer, Norton, Allen, & Steele, 1973) for high or middle autism as compared to the uptake in the presence of serum from nonpsychotic children. Cohen, Caparulo, Shaywitz, and Bowers (1977) measured cerebrospinal fluid (CSF) levels of 5-hydroxyindolacetic acid (5-HIAA), a metabolite of serotonin, and of homovanillic acid (HVR), a metabolite of dopamine, after probenecid blockade. They compared findings for a group of severely autistic children to findings in four other groups of children: (1) nonautistic psychoses of early childhood children, (2) central processing disturbance children, (3) aphasic children, and (4) children being evaluated pediatrically for suspected CNS disorder. There was no significant difference between the autistic children for CSF levels of 5-HIAA or HVA and group 2, 3, or 4 of the other children. The autistic children did have significantly lower 5-HIAA levels than the nonautistic psychosis of early childhood children. In another article Cohen and Young (1977) reported that some autistic children with lowered 5-HIAA CSF levels also had elevated HVA CSF levels. These children appeared to be the most deviant clinically with increased stereotypy and hyperactivity. They were also able to measure the CSF 5-HIAA and HVA levels in a set of monozygotic twins concordant for autism. One twin was consistently more hyperactive and engaged in more stereotypic behavior (flapping and twirling) than the other. His CSF HVA level was higher than that of his twin. Cohen and Young felt that these findings may indicate that in autistic and other psychotic children increased dopamine turnover may be associated with increased activity and abnormal movement patterns. They suggest that it may be of more value to look for biologic correlates of specific symptoms than to look for syndrome-specific biologic correlates.

Tryptophan Loading Studies In 1958, Sutton and Read reported a case of an autistic child who on receiving .25 mg/kg of L-tryptophan secreted less 5-HIAA in his urine than did four nonpsychotic hospitalized children who had ingested a similar amount of L-tryptophan. They felt this might indicate that autistic children might have a diminished capacity to metabolize tryptophan via the serotonin chain. Shaw, Lucas, and Rabinovitch (1959) were unable to duplicate the finding in schizophrenic as compared to nonschizophrenic children but pointed out that this didn't mean there was no difference in brain serotonin metabolism in schizophrenic as compared to nonschizophrenic children, since only a small amount of orally ingested tryptophan is metabolized through the 5 hydroxyindole pathway and of this only a small portion is metabolized in the brain. Schain and Freedman (1961) also found no differ-

Piggott

210

ences in urinary excretion of 5-HIAA after tryptophan loading in autistic as compared to mildly retarded nonautistic as compared to severely retarded nonautistic children, but they did find that the autistic and severely retarded children had higher blood serotonin levels than did the mildly retarded group. Within the autistic group (N = 23) there were six children who had persistently very high blood serotonin levels. Clinically they were indistinguishable from the other autistic children. In 1977 Hanley, Stahl, and Freedman reported on extensions of the Schain and Freedman study. They compared groups of autistic, severely retarded nonautistic, mildly retarded nonautistic, and normal children to each other for blood serotonin levels. They found hyperserotonemia in 30%, 52o70, and 9%, respectively, of the first three groups but no hyperserotonemia in the normal control children. Next, they measured the urinary excretion of serotonin, 5-HIAA, tryptamine, and vanillylmandelic acid (VMA) in four hyperserotonemic autistic children and four normoserotonemic mildly retarded children. The autistic children excreted more of all four metabolites. Both groups were then given L-tryptophan 3 gm per day for 3 days. This resulted in a marked increase in 5-HIAA excretion in both groups and an increase in urinary serotonin excretion in the hyperserotonemic autistic children but the opposite effect (a decrease in urinary serotonin excretion) in the mildly retarded children. Thus it seems that there may be differences in urinary, blood, and spinal fluid levels of components of the serotonin metabolic chain in autistic as compared to other children. Yet these differences may not be unique to autistic children since some have been shown to be shared by severely retarded children. To what degree they may be shared by other groups of developmentally deviant children is untested and whether any of them are etiologically related to autism is far from clear. Platelet Serotonin Level Studies

Ritvo et al. (1970) found an inverse relationship between age and blood serotonin levels and between age and platelet count in normal children but not in autistic children. In autistics they also found Significantly higher blood serotonin levels and significantly higher blood platelet counts than in the normal children. Takahashi, Kanai, and Miyamoto (1976) compared 30 autistic to 30 normal and 30 nonautistic children with various neurological and psychiatric disorders for platelet serotonin levels. They found that the autistic children as a group had a mean platelet serotonin level significantly higher than the mean of the normal group. However, one-third of the autistic children had platelet serotonin levels below the mean of the normal children. The mean platelet serotonin level of the autistic children did not differ significantly from that of the nonautistic

Overview of Research in Autism

211

neurologically and psychiatrically disturbed children. They rated the autistic and nonautistic disturbed children for hyperactivity and found that the hyperactive children tended to have higher platelet serotonin levels than the nonhyperactive children. Additionally, they examined platelets from 34 parents of autistic children and found marked variation in platelet serotonin levels, but though higher they were not significantly higher than serotonin levels in platelets from healthy adults without autistic children. Takahashi et al. concluded that elevated platelet serotonin levels do occur in infantile autism but may not be specific to infantile autism. They pointed out that factors such as diet and activity levels had not been systematically studied so that one cannot state that their findings truly represent a biologic difference in the autistic as compared to the nonautistic child. One can speculate on the lack of significant difference in the autistic group and the nonautistic disturbed group, the wide variation in levels, and the tendency for the higher levels to be found in the hyperactive members of both groups. What is the overlap or what developmental deviations may both groups have in common?

Platelet Uptake and Efflux of Serotonin Studies Blood platelets and serotonin-containing neurons both have active serotonin transport systems and store serotonin in subcellular organelles of similar electron microscopic appearance. In 1962, Sankar, Gold, Phipps, and Sankar reported that the uptake of serotonin by platelets from schizophrenic children is lower than that by platelets from nonpsychotic hospitalized children. Boullin, Coleman, and O'Brien (1970) found a twofold increase in platelet serotonin efflux in autistic children diagnosed by Rimland's criteria as compared to a group of comparably aged controls. Lucas, Warner, and Gottlieb (1971) found no significant differences in platelet serotonin uptake in a group of schizophrenic children versus a group of nonpsychotic emotionally disturbed children. Boullin, Coleman, O'Brien, and Rimland (1971), by using a threefold increase in platelet serotonin efflux as a marker, correctly diagnosed six of seven autistic children who scored over 20 on the Rimland E-2 scale. They also found a clear increase in platelet serotonin uptake in autistic children as compared to controls. They suggested that the increased serotonin efflux might be due to defective serotonin binding by the platelet and this defective binding may also be present in the neuron. Yuwiler, Ritvo, Geller, Glousman, Schneiderman, and Matsuno (1975), in attempting to repeat the findings of BouUin et al. of increased uptake and efflux of serotonin by the platelets, found no differences among autistic children, nonautistic psychiatrically hospitalized children, and normals. They used two separate procedures to make their

212

Piggott

platelet serotonin uptake and efflux measurements. Rimland (1976) felt their inability to duplicate the findings was because they used a different group of autistic children. Their group was diagnosed by criteria in which perceptual inconstancy is an integral part, whereas the group used by Boullin et al. was diagnosed by Rimland's checklist on which only 10~ of psychotic children score in the autistic range and in which the symptom "insistence on the preservation of sameness" is an integral part. Rimland then stated that since the groups of autistic children used in the two studies were basically different it was to be expected that the results would be different. At this point it appears that there may be an increase in platelet serotonin efflux in some autistic children diagnosed by the Rimland E-2 scale. Autistic children not diagnosed by that scale have not been shown to have any consistent increase in either platelet serotonin efflux or uptake as compared to other children. It may well be that children diagnosed as autistic by the Rimland E-2 scale represent a subgroup of autistic children who are basically different from other autistic children. However, perceptual inconstancy is not necessarily incompatible with the symptom insistence on sameness in that an attempt to cope with inner inconstancy might well involve trying to maintain environmental constancy (preservation of sameness).

Enzyme Studies Enzyme studies have been carried out by a few investigators. Boullin, Bhagavan, Coleman, O'Brien, and Youdim (1975), Campbell, Friedman, Green, Small, and Burdock (1976), Lake et al. (1977), and Cohen, Young, and Roth (1977) have all reported negative findings for platelet MAO activity differences in autistic or schizophrenic children as compared to nonpsychiatrically disturbed controls. Katz and Liebman (1970) reported elevated CSF creatine phosphokinase (CPK) activity in 3 of 4 autistic children. They also found elevation of CSF CPK activity in 8 of 17 patients with bacterial meningitis, 6 of 14 patients with nonbacterial meningitis, 4 of 10 patients with febrile convulsions, and 10 of 34 patients with epilepsy. This may be evidence for CNS damage in autism. Sankar (1971) reported significantly lower blood adenosine triphosphatase (ATPase) activity in assays of rbc's from autistic children when the blood cells remained intact but not when the rbc's were lysed. He believes these results indicate that the rbc membranes of autistic children display a decreased permeability to ATP or to some factors necessary for ATPase activity in the cell. These studies merit further exploration. Autistic children with increased CSF CPK activity may represent a subgroup of children whose autism is due to brain insult from infection and Sankar's findings, if duplicated, may represent a different subgrouping of autistic children.

Overview of Research in Autism

213

Immunologic Studies Immunologic studies indicate there may be an immunologic system deficiency in some autistic children. Fowle (1968) found an increased frequency of proplasmocytes, cells that have been shown to be associated with allergies and viral infections, in schizophrenic children as compared to their siblings and a group of unrelated normal children. Stubbs (1976) found undetectable hemagglutination-inhibition antibody titers in some autistic children following rubella vaccination. In 1977 he and his colleagues (Stubbs, Crawford, Burger, & Vandenbark) reported finding a significantly depressed lymphocytic responsiveness in a group of autistic as compared to normal children. All the autistic children were retarded, one was deaf and cerebral palsied due to cytomegalovirus. Young, Caparulo, Shaywitz, Johnson, and Cohen (1977) found no evidence of elevated CSF immunoglobulin levels in 15 autistic children. The possibility of a defective immunologic response interacting with an environmental agent to affect brain development possibly during critical periods, thus resulting in relatedness, communicative, and other disturbances seen in autism and perhaps in other types of developmental deviations, is intriguing and well worth further study. In this context Folstein and Rutter's (1977) report on a study of 21 twin pairs in which at least 1 twin had early onset childhood psychosis is of interest. Ten of the 21 twin pairs were dizygotic, the other 11 pairs were monozygotic. Four (36~ of the monozygotic co-twins were concordant for autism, none of the dizygotic co-twins were concordant for autism. Nine (82~ of the monozygotic co-twins were also concordant for cognitive impairment, whereas only 1 (10~ of the dizygotic co-twins were concordant for cognitive disturbance. Eleven of the 42 children (21 twin pairs) had a history of biological hazards associated with the birth process. This finding did not account for the concordance patterns. However, in the 6 pairs that were discordant for autism and had a history of birth process hazards, it was the autistic twin who had suffered the biologic hazard. Rutter believes this study clearly demonstrates the importance of genetic factors in earlyonset autism in addition to indicating that autism is also associated with biological hazards in infancy.

DISCUSSION There is accumulating evidence that organic disturbances are related to and may be responsible for many of the symptoms seen in various children called autistic. Which and to what degree these disturbances are unique to autism or are also involved in a variety of other developmental deviations is unclear and in need of continued research. From an etiologic

214

Piggott

point of view, the children called autistic probably are composed of a number of subgroups with different basic pathologic processes manifesting in similar clinical pictures. There is increasing indication that at least some of the disturbances originate during gestation, are present at birth, and become manifest when they interfere with the normal course of development. The idea of genetic vulnerability combined with environmental insult to produce autism, as suggested by Folstein and Rutter (1977), seems likely to account for some autistic children. The idea of Stubbs et al. of an immunologic deficit followed by a viral insuk would fit this pattern. The first trimester of pregnancy might be a time of particular susceptibility, as suggested by Steg and Rapoport (1975) from their findings of increased minor anomalies in autistic children. The possibility of a basic disturbance affecting the body's feedback systems, as suggested by Goldfarb's (1974) findings, the free fatty acid studies of DeMyer et al. (1971), the autonomic studies, and the vestibular studies, particularly those of the UCLA group, or of a basic disturbance affecting cell membrane function, as suggested by Sankar's (1971) ATPase studies, the findings on platelet serotonin efflux of Boullin et al. (1971), and the tryptophan uptake study of Piggott et al. (1975), may be an explanation whereby one basic dysfunction could result in a multiplicity of variable symptoms. We are in need of objective markers whereby various subgroups of children diagnosed as autistic or schizophrenic can be clearly differentiated so as to facilitate thorough study toward a clarification of u n d e r l y i n g pathology. This, then, could lead toward specific effective therapy. Perhaps the decrease or absence of postrotatory nystagmus under lighted conditions, as demonstrated by Ritvo et al. (1969), the auditory evoked response by which Saletu et al. (1975) could differentiate high risk from control children, the decreased lymphocyte responsiveness found by Stubbs et al. (1977), the presence of urinary bufotenin and/or DMT demonstrated by Narasimhachari and Himwich (1975), or the increased blood platelet serotonin efflux by which Boullin et al. (1971) accurately differentiated six of seven autistic children diagnosed by the Rimland E-2 scale may, if they can be duplicated by other investigators, become markers for subgroups of autistic children. We also would do well to heed Cohen and Young's (1977) suggestion of focusing on correlating biological findings with specific symptoms rather than with constellations of symptoms that tend to confuse us.

REFERENCES

Bernal, M. E., & Miller,W. H. Electrodermaland cardiac responsesof schizophrenicchildren to sensorystimuli. Psychophysiology, 1970, 7, 155-168. Boullin, D. J., Bhagavan,H. N., Coleman, M., O'Brien, R. A., & Youdim, M. B. H. Platelet monoamir, oxidase in children with infantile autism. Medical Biology, 1975, 53, 210-213.

Overview of Research in Autism

215

Boullin, D. J., Coleman, M., & O'Brien, R. A. Abnormalities in platelet 5-hydroxytryptamine efflux in patients with infantile autism. Nature, 1970, 226, 371-372. Boullin, D. J., Coleman, M., O'Brien, R. A., & Rimland, B. Laboratory predictions of infantile autism based on 5-hydroxytryptamine efflux from blood platelets and their correlation with the Rimland E-2 score. Journal of Autism and Childhood Schizophrenia, 1971, 1, 63-71. Campbell, M., Friedman, E., DeVito, E., Greenspan, L., & Collins, P. Blood serotonin in psychotic and brain damaged children. Journal of Autism and Childhood Schizophrenia, 1974, 4, 33-41. Campbell, M., Friedma n, E., Green, W. H., Collins, P. J., Small, A. M., & Breuer, H. Blood serotonin in schizophrenic children. A preliminary study. International Pharmacopsychiatry, 1975, 10, 213-221. Campbell, M., Friedman, E., Green, W. H., Small, A. M., & Burdock, E. I. Blood platelet monoamine oxidase activity in schizophrenic children and their families. Neuropsychobiology, 1976, 2, 239-246. Cohen, D. J., Caparulo, B. K., Shaywitz, B. A., & Bowers, M. B. Dopamine and serotonin metabolism in neuropsychiatrically disturbed children. Archives of General Psychiatry, 1977, 34, 545-550. Cohen, D., & Young, J. G. Neurochemistry and child psychiatry. Journal of ChiM Psychiatry, 1977, 16, 353-411. Cohen, D., Young, J. G., & Roth, J. A. Platelet monoamine oxidase in early childhood 9 autism. Archives of General Psychiatry, 1977, 34, 534-537. Colbert, E. G., Koegler, R. R., & Markham, C. H. Vestibular dysfunction in childhood schizophrenia. Archives of General Psychiatry, 1959, 1, 600-617. Coleman, M., Campbell, M , Freedman, L. S., Roffman, M., Ebstein, R. P., & Goldstein, M. Serum dopamine-/3-hydroxylase levels in Down's syndrome. Clinical Genetics, 1974, 5, 312-315. Creak, M., & Pampiglione, G. Clinical and EEG studies on a group of 35 psychotic children. Developmental Medicine and Child Neurology, 1969, 11, 218-227. Darby, J. K. Neuropathologic aspects of psychosis in children. Journal of Autism and Childhood Schizophrenia, 1976, 6, 339-352. DeMyer, M., Barton, S., DeMyer, W., Norton, J., Allen, J., & Steele, R. Prognosis in autism: A follow-up study. Journal of Autism and Childhood Schizophrenia, 1973, 3, 199-246. DeMyer, M., Schwier, H., Bryson, C., Solow, E., & Roeske, N. Free fatty acid response to insulin and glucose stimulation in schizophrenic, autistic, and emotionally disturbed children. Journal of Autism and Childhood Schizophrenia, 1971, 1, 436-452. Fish, B. Biologic antecedents of psychosis in children. In D. X. Freedman (Ed.), Biology of the major psychoses. New York: Raven Press, 1975. Folstein, S., & Rutter, M. Infantile autism: A genetic study of 21 twin pairs. Journal of Child Psychology and Psychiatry, 1977, 18, 297-321. Fowle, A. M. Atypical leukocyte pattern of schizophrenic children. Archives of General Psychiatry, t968, 18, 666-680. Freeman, B. J., Frankel, F., & Ritvo, E. R. The effects of response contingent vestibular stimulation on the behavior of autistic and retarded children. Journal of Autism and Childhood Schizophrenia, 1976, 6, 353-358. Goldfarb, W. Factors in the development of schizophrenic children: An approaela to subclassification. In J. Romano (Ed.), The origins of schizophrenia. Amsterdam: Excerpta Medica Foundation, 1967. Pp. 70-91. Goldfarb, W. Growth and change of schizophrenic children: A longitudinal study. New York: Wiley, 1974. Goldstein, M., Mahanand, D., Lee, J., & Coleman, M. Dopamine-/3-hydroxylase and endogenous total 5-hydroxyindole levels in autistic patients and controls. In M. Coleman (Ed.), The autistic syndromes. New York: American Elsevier, 1976. Gubbay, S. S., Lobascher, M., & Kingerlee, P. A neurological appraisal of autistic children: Results of a Western Australian survey. DevelopmentalMedicine and ChiM Neurology, 1970, 12, 422-429. Hanley, H. G., Stahl, S. M., & Freedman, D. X. Hyperserotonemia and amine metabolites in autistic and retarded children. Archives of General Psychiatry, 1977, 34, 521-531.

216

Piggott

Hauser, S., Delong, G., & Rosman, N. Pneumographic findings in the infantile autism syndrome: A correlation with temporal lobe disease. Brain, 1975, 98, 667-688. Hurt, C., Forrest, S. J., & Richer, J. Cardiac arrhythmia and behavior in autistic children. Acta Psychiatrica Scandinavica, 1975, 51, 361-372. Hutt, S. J., Hutt, C., Lee, D., & Ounsted, C. A behavioural and electroencephalographic study of autistic children. Journal of Psychiatric Research, 1965, 3, 181-197. :leannerod, M., Mouret, J., & Jouvet', M. Etude de la motricit6 oculaire au cours de la phase paradoxale du sommeil chez le chat. Electroencephalography and Clinical Neurophysiology, 1965, 18, 554-566. Katz, R. M., & Liebman, W. Creatine phosphokinase activity in central nervous system disorders and infections. American Journal of Diseases of Children, 1970, 120, 543-546. Lake, C. R., Ziegler, M. G., & Murphy, D. L. Increased norepinephrine levels and decreased dopamine-/J-hydroxylase activity in primary autism. Archives of General Psychiatry, 1977, 34, 553-556. Lelord, G., Laff0nt, F., Jusseaume, P., & Stephant, J. L. Comparative study of conditioning of averaged evoked responses by coupling sound and light in normal and autistic children. Psychophysiology, 1973, 10, 415-425. Lucas, A. R., Warner, K., & Gottlieb, J. S. Biological studies in childhood schizophrenia: Serotonin uptake by platelets. Biological Psychiatry, 1971, 3, 123-128. Magherini, P. C., Pompeiano, O., & Thoden, U. The neurochemical basis of REM sleep: A cholinergic mechanism responsible for'rhythmic activation of the vestibulo-oculomotor system. Brain Research, 1971, 35, 565-569. Medni'ck, S. A., & Schulsinger, F. Some premorbid characteristics related to breakdown in children with schizophrenic mothers. In D. Rosenthal and S. S. Kety, The transmission of schizophrenia. Oxford: Pergamon Press, 1968. Pp. 267-291. I Narasimhachari, N., & Himwich, H. E. Biochemical studies in early infantile autism. Biological Psychiatry, 1975, 10, 425-432. Ornitz, E. M., Brown, M. B., Mason, A., & Putnam, N. H. Effect of visual input on vestibular nystagmus in autistic children. Archives of General Psychiatry, 1974, 31, 369-375. Ornitz, E. M., Ritvo, E. R., Brown, M. B., La Franchi, S., Parmelee, T., & Walter, R. D. The General Psychiatry, 1968, 18, 76-98. Ornitz, E. M., Ritvo, E. R., Brown, M. B., La Franchi, S., Parmelee, T., & Walter, R. O. The EEG and rapid eye movements during REM sleep in normal and autistic children. Electroencephalography and Clinical Neurophysiology, 1969, 26, 167-175. Ornitz, E. M., Ritvo, E. R., Panman, L. M., Lee, Y. H., Carr, E. M., & Waiter, R. D. The auditory evoked response in normal and autistic children during sleep. Electroencephalography and Clinical Neurophysiology, 1968, 25, 221-230. Pare, C. M. B., Sandler, M., & Stacey, R. S. The relationship between decreased 5-hydroxyindole metabolism and mental defect in phenylketonuria. Archives of Disease in Childhood, 1959, 34, 422-425. Partington, M. W., Tu, J. B., & Wong, C. Y. Blood serotonin levels in severe mental retardation. Developmental Medicine and Child Neurology, 1973, 15, 616-627. Perry, T. L. N-methylmetanephrine excretion by juvenile psychotics. Science, 1963, 139, 587-589. Piggott, L. R., Ax, A. F., Bamford, J. L., & Fetzner, J. M. Respiration sinus arrhythmia in psychotic children. Psychophysiology, 1973, 10, 401-414. Piggott, L. R., Frohman, C. E., Ward, V. L., & Gottlieb, J. S. The effect of plasma from psychotic children on tryptophan uptake in chicken erythrocytes. Neuropsychobiology, 1975, 1, 284-295. Piggott, L. R., Purcell, G., Cummings, G., & Caldwell, D. Vestibular dysfunction in emotionally disturbed children. Biological Psychiatry, 1976, 11, 719-729. Pollack, M., & Krieger, H. P.'Oculomotor and postural patterns in schizophrenic children. A.M.A. Archives of Neurology and Psychiatry, 1958, 79, 720-726. Quinn, P., & Rapoport, J. Minor physical anomalies and neurologic status in hyperactive boys. Pediatrics, 1974, 53, 742-747. Rimland, B. Platelet uptake and efflux of serotonin in subtypes of psychotic children. Journal of Autism and Childhood Schizophrenia, 1976, 6, 379-382.

Overview of Research in Autism

217

Ritvo, E. R., Ornitz, E. M., Eviatar, A., Markham, C. H., Brown, M. B., & Mason, A. Decreased postrotatory nystagmus in early infantile autism. Neurology, 1969, 19, 653-658. Ritvo, E. R., Omitz, E. M., Walter, R. D., & Hanley, J. Correlation of psychiatric diagnoses and EEG findings: A double-blind study of 184 hospitalized children. American Journal of Psychiatry, 1970, 126, 988-996. Ritvo, E. R., Yuwiler, A., Geller, E., Ornitz, E. M., Saeger, K., & Plotkin, S. Increased blood serotonin and platelets in early infantile autism. Archives of General Psychiatry, 1970, 23, 566-572. Saletu, B., Saletu, M., Marasa, J., Mednick, S., & Schulsinger, F. Acoustic evoked potentials in offspring of schizophrenic mothers. Clinical Electroencephalography, 1975, 6, 92-102. Sankar, D. V. S. Studies on blood platelets, blood enzymes, and leukocyte chromosome breakage in childhood schizophrenia. Behavioral Neuropsychiatry, 1971, 2, 2-10. Sankar, D. V. S., Gold, E., Phipps, E., & Sankar, D. B. General metabolic studies on schizophrenic children. Annals of the New York Academy of Sciences, 1962, 96, 392397. Schain, R. J., & Freedman, D. X. Studies on 5-hydroxyindole metabolism in autistic and other mentally retarded children. Journal of Pediatrics, 1961, 58, 315-320. Shaw, C., Lucas, J., & Rabinovitch, R. Metabolic studies in childhood schizophrenia--effects of tryptophan loading on indole excretion. Archives of General Psychiatry, 1959, I, 366-371. Small, J. G. Sensory evoked responses of autistic children. In D. W. Churchill, G. D. Alpern, & M. DeMyer (Eds.), Infantile autism: Proceedings of the Indiana University Colloquium. Springfield, Illinois: Charles C Thomas, 1971. Pp. 224-242. Small, J. G. EEG and neurophysiological studies of early infantile autism. Biological Psychiatry, 1975, 10, 385-397. Steg, J. P., & Rapoport, J. L. Minor physical anomalies in normal, neurotic, learning disabled, and severely disturbed children. Journal of Autism and Childhood Schizophrenia, 1975, 5, 299-307. Stubbs, E. G. Autistic children exhibit Undetectable hemagglutination-inhibition antibody titers despite previous rubella vaccination. Journal of Autism and Childhood Schizophrenia, 1976, 6, 269-274. Stubbs, E. G., Crawford, M., Burger, D. R., & Vandenbark, A. A. Depressed lymphocyte responsiveness in autistic children. Journal of Autism and Childhood Schizophrenia, 1977, 7, 49-55. Student, M., & Sohmer, H. Evidence from auditory nerve and brainstem evoked responses for an organic brain lesion in children with autistic traits. Journal of Autism and Childhood Schizophrenia, 1978, 8, 13-20. Sutton, H. E., & Read, J. Abnormal amino acid metabolism in a case suggesting autism. American Journal of Diseases of Children, 1958, 98, 23-25. Takahashi, S., Kanai, H., & Miyamoto, Y. Reassessment of elevated serotonin levels in blood platelets in early infantile autism. Journal of Autism and Childhood Schizophrenia, 1976, 6, 317-326. Tanguay, P. E. Clinical and electrophysiological research. In E. R. Ritvo (Ed.), Autism: Diagnosis, current research and management. Holliswood, New York: Spectrum Publications, 1976. Tanguay, P. E., Ornitz, E. M., Forsythe, A. B., & Ritvo, E. R. Rapid eye movement (REM) activity in normal and autistic children during REM sleep. Journal of Autism and Childhood Schizophrenia, 1976, 6, 275-288. Tu, J. B., & Partington, M. W. 5-bydroxyindole levels in the blood and CSF in Down's syndrome, phenylketonuria, and severe mental retardation. Developmental Medicine and Child Neurology, 1972, 14, 457-466. , Waldrop, M., Pedersen, F. A., & Bell, R. Q. Minor physical anomalies and behavior in preschool children. Child Development, 1968, 39, 391-400. White, P., DeMyer, W., & DeMyer, M. EEG abnormalities in early childhood schizophrenia: A double-blind study of psychiatrically disturbed and normal children during promazine sedation. American Journal of Psychiatry, 1964, 120, 950-958.

218

Piggott

Young, J. G., Caparulo, B., Shaywitz, B., Johnson, W., & Cohen, D. Childhood autism: Cerebrospinal fluid examination and immunoglobulin levels. Journal of Child Psychiatry, 1977, 16, 174-179. Yuwiler, A., Ritvo, E., Geller, E., Glousman, R., Schneiderman, G., & Matsuno, D. Uptake and efflux of serotonin from platelets of autistic and nonautistic children. Journal of Autism and Childhood Schizophrenia, 1975, 5, 83-98.