TONG, ROSS, SCHMUNK, ET AL. have yet been provie inicating that opamine D 1 receptor agonists are rewaring to humans (14), agonists acting on this rec

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1 Article Decrease Striatal Dopamine D 1 Receptor- Stimulate Aenylyl Cyclase Activity in Human Methamphetamine Users Junchao Tong, Ph.D. Brian M. Ross, Ph.D. Gregory A. Schmunk, M.D. Frank J. Peretti, M.D. Kathryn S. Kalasinsky, Ph.D. Yoshiaki Furukawa, M.D. Lee Cyn Ang, M.D. Sally S. Aiken, M.D. Dennis J. Wickham, M.D. Stephen J. Kish, Ph.D. Objective: It has been assume that some behavioral changes associate with repeate exposure to opaminergic psychostimulant rugs might be explaine by changes in activity of opamine receptors, incluing the opamine D 1 receptor, which is linke by a stimulatory G protein to the effector enzyme aenylyl cyclase. To establish whether opamine D 1 receptor function might be altere in human methamphetamine users, the authors measure opamine-stimulate aenylyl cyclase activity in the brain of chronic human users of the rug. Metho: Aenylyl cyclase activity stimulate by opamine an by guanylyl-imioiphosphate (to assess G protein an aenylyl cyclase coupling) was etermine in the postmortem brain tissue of 16 methamphetamine users who ha use the rug both recently an chronically (i.e., at least 1 year) as well as 21 matche comparison subjects. Results: A 25% 3% ecrease in the maximal extent of opamine stimulation of aenylyl cyclase activity was seen in the striatum (nucleus accumbens, cauate, an putamen) of the methamphetamine users. No changes were foun in basal or guanylyl-imioiphosphate-stimulate enzyme activity. Conclusions: These ata suggest that opamine receptor function linke to aenylyl cyclase is partially esensitize in the striatum of human methamphetamine users. Decrease opamine D 1 receptor function might unerlie part of the known (rug withrawal synrome) or expecte (rug tolerance) consequences of methamphetamine exposure in humans. (Am J Psychiatry 23; 16:896 93) Methamphetamine is a highly aictive psychostimulant that is wiely abuse for the euphoric effects of the rug. The chronic effects of methamphetamine exposure in some, but not all, regular rug users inclue an extene rug withrawal synrome that can be characterize by separate perios of ysphoria an rug craving (1). It is also assume that with repeate use of methamphetamine, some tolerance usually occurs to the pleasurable effects of the rug (2). It is surprising, however, that the question of acute or chronic tolerance has not yet been rigorously establishe in chronic human users of the rug (3). Methamphetamine shares with other rugs of abuse (e.g., cocaine, heroin, alcohol) the ability to enhance release of the neurotransmitter opamine in the striatum (cauate, putamen, nucleus accumbens) (4, 5), with the extent of the opamine increase correlate with the intensity of the high in humans (6, 7). For this reason, it continues to be assume that some of the effects of methamphetamine exposure in humans are explaine in large part by alterations in activity of the pre- an postsynaptic opamine system in the brain (8). Thus, we previously reporte that striatal tissue levels of opamine can be low in some users of methamphetamine (9) as low as those in Parkinson s isease in some striatal subivisions (1) which suggests that some of the unpleasant consequences of methamphetamine uring rug withrawal (e.g., epression, cognitive impairment) might be ue in part to a striatal opamine eficiency. It is also possible that shortan long-term aaptive changes ue to overactivation of opamine receptors occurs in methamphetamine-expose human brain, which might explain some aspects of rug-taking behavior. Dopamine receptor types can be classifie by their ability to stimulate (D 1 receptors) or inhibit (D 2 receptors) aenylyl cyclase (EC ) through the meiation of the stimulatory G protein (G s or, in striatum, G olf [11]) or the inhibitory G protein (G i, G o ), respectively (see reference 12 for a review). Since the pharmacological literature on the influence of opamine D 1 receptor agonists/antagonists on the behavior of human psychostimulant users is scanty, the role of the opamine D 1 receptor in psychostimulant use an abuse in the human has not yet been establishe. However, the preliminary observation that a single ose of a opamine D 1 receptor antagonist (SCH 39166, ecopipam) can partially block the euphoric effects of cocaine in chronic users of the psychostimulant (13) suggests that the opamine D 1 receptor might meiate part of the pleasurable effects of opaminergic stimulants. Although no ata Am J Psychiatry 16:5, May 23

2 TONG, ROSS, SCHMUNK, ET AL. have yet been provie inicating that opamine D 1 receptor agonists are rewaring to humans (14), agonists acting on this receptor are self-aministere by nonhuman primates (15, 16). Apart from these pharmacological stuies, information on the status of the opamine D 1 receptor system in human psychostimulant users is limite to our postmortem fining of normal levels of opamine D 1 receptor protein in the opamine-rich striatal subivisions of methamphetamine users an users of cocaine, with the exception of higher protein concentration of the receptor restricte to the nucleus accumbens subivision in the methamphetamine users (17). Changes or lack of changes in brain neurotransmitter receptor concentrations can, at most, only suggest ifferences in receptor function. Therefore, we have now extene our investigation to inclue measurement, in the postmortem brain tissue of chronic methamphetamine users, of a more ynamic inex of opamine D 1 receptor function, namely opamine-stimulate aenylyl cyclase activity, since aenylyl cyclase is consiere to be the key effector of opamine D 1 receptor function (12). In orer to establish the possible site an specificity of any isturbance in opamine D 1 receptor, G protein, an aenylyl cyclase coupling, we also measure stimulation of aenylyl cyclase by a nonhyrolyzable GTP analog guanylyl-imioiphosphate [Gpp(NH)p], which stimulates aenylyl cyclase by irect activation of the stimulatory G protein, i.e., bypassing the opamine receptor, thereby assessing the G protein an aenylyl cyclase coupling. We report that opamine D 1 -stimulate aenylyl cyclase activity is ecrease in striatum of human methamphetamine users, a fining that might explain some of the short- or longterm aspects of rug-taking behavior. Metho Brain Materials Brains from a total of 21 comparison subjects (19 men an two women) an 16 chronic users of methamphetamine (11 men an five women) were obtaine postmortem from meical examiner offices in the Unite States per a stanarize protocol. Informe consent was obtaine from the next of kin. The comparison subjects an methamphetamine users i not significantly iffer in age (mean=33.6 years [SD=1.1] an 32.4 years [SD=8.1], respectively; t=.38, f=35, p=.7) or postmortem interval between eath an freezing of the brain (mean=13.1 hours [SD=5.8] an 15.3 hours [SD=6.7]; t=1.1, f=35, p=.29). At autopsy, half of the brain was fixe in formalin fixative for neuropathological analysis, whereas the other half was immeiately frozen until issection for neurochemical analysis. Samples of femoral bloo were obtaine from all of the methamphetamine users an the comparison subjects for rug screening. Scalp hair samples for rug analyses were available for 18 of the 21 comparison subjects an 12 of the 16 methamphetamine users. Levels of rugs of abuse in bloo an other boily fluis were measure by the local meical examiner; rug analyses in brain an hair samples were conucte by one of the investigtors (K.K.) at the Arme Forces Institute of Pathology in Washington, D.C. All methamphetamine users met the following selection criteria: 1) presence of methamphetamine confirme by toxicology screening analyses of bloo, autopsie brain, an, when available, sequential scalp hair samples; 2) absence of any other rugs of abuse (incluing bloo ethanol) in these tissues; 3) evience of methamphetamine use for at least 1 year before eath (obtaine from case recors an structure interviews with meical examiner investigators, next of kin, an informants); an 4) absence of neurological illness or brain pathology unrelate to use of the rug. Drug histories an patient information are summarize in Table 1, with information on 12 of the 16 methamphetamine users previously reporte (9). The regional istribution of methamphetamine an its metabolite amphetamine in autopsie brain has been reporte for 12 of the 16 methamphetamine subjects (18). All comparison subjects were neurologically normal, ha no evience of brain pathology, ha no history of rug use, an teste negative for all rugs of abuse in bloo, autopsie brain, an, in the 18 subjects from whom it was available, sequential scalp hair samples. The causes of eath for the comparison subjects were cariovascular isease (N=12), trauma (N=7), rowning (N=1), an leukemia (N=1). Aenylyl Cyclase Assay Homogenates, in 5 mm Tris-HCl, ph 7.4,.1 mm CaCl 2, 1% (vol/vol) protease inhibitors (Sigma, catalog number P834), of issecte brain samples (nucleus accumbens, cauate, putamen, frontal cortex [Bromann s area 9], an temporal cortex [Bromann s area 22]) were use. The proceure for the assay of opamine-stimulate aenylyl cyclase activity was the same as reporte previously (19) with minor moifications. Enzyme activity in brain homogenates (2 µg protein) was assaye in a total volume of 1 µl containing 5 mm Tris-HCl buffer, ph 7.4,.5 mm MgCl 2,.2 mm EGTA, 4 mm phosphocreatine, 25 units/ml of creatine phosphokinase, 1% (vol/vol) protease inhibitors, 1 mm camp, 1 µm GTP,.2 mm ATP, an 1 µci [α- 33 P]ATP in the absence an presence of varying concentrations (.4 4 µm, seven concentrations) of opamine. In the present stuy, the concentration of Mg ++ was lowere to.5 mm (versus 4 mm in reference 19) in the assay, since this moification prouce lower basal aenylyl cyclase activity an therefore higher percentage opamine stimulation in the striatal tissues (3% 4% versus approximately 2%). The percentage stimulation in the cerebral cortex was not change by the moification. For the assay of Gpp(NH)p stimulation, GTP was replace with varying concentrations (1 8 to 1 4 M, five concentrations) of Gpp(NH)p, an the concentration for MgCl 2 was 5 mm. Uner the conitions use, i.e., high ATP concentration, high temperature (3 C), an without an aitional factor (e.g., forskolin, calcium/calmoulin) to elevate basal aenylyl cyclase activity, Gpp(NH)p preferentially activates G s /G olf rather than G i /G o an thus stimulates rather than inhibits aenylyl cyclase (Tong an Kish, unpublishe results; reference 2). Samples were first pre-incubate with the assay mixture on ice for 1 minutes an the reaction initiate by aition of ATP. The assay was carrie out at 3 C for 3 minutes (opamine stimulation) or 2 minutes (Gpp[NH]p stimulation). The assay was terminate by the aition of 2% soium oecylsulfate, 4 mm ATP, an 1.4 mm camp, followe by boiling for 1 minutes. Aenylyl cyclase activity was etermine by the Dowex/Alumina twocolumn chromatography proceure of Salomon et al. (21) as escribe (19). Assays were carrie out in uplicate, an activity was expresse as pmol camp forme per minute per mg of protein. Protein concentration was etermine by using the Bio-Ra Protein Assay Kit with bovine plasma albumin as the stanar. Basal aenylyl cyclase activity was efine as the activity in the absence of any aitional factors. Basal aenylyl cyclase with GTP was the activity in the presence of 1 µm GTP an was use for calculating percentage opamine stimulation. The presence of GTP was require for the stimulation by opamine an the con- Am J Psychiatry 16:5, May

3 DOPAMINE RECEPTORS AND METHAMPHETAMINE TABLE 1. Characteristics an Drug Use History of 16 Methamphetamine Users a Toxicology Confirmation of Methamphetamine Use Subject Age (years) Sex Postmortem Interval (hours) Duration of Use (years) Recent Drug Use Pattern Route of Aministration Suspecte/Known Cause of Death Hair Sample Brain Drug Level b 1 34 F 14 1 Daily Nasal Methamphetamine M 5 >1 Once per Nasal; Methamphetamine 8.4 month intravenous 3 22 M 16 8 Daily, limite Intravenous Methamphetamine 12.9 only by funs 4 42 M 1 >2 3 4 times per Nasal; oral Methamphetamine 12.9 week 5 2 M 21 1 Unknown Oral Methamphetamine Yes M Daily an 2 3- Intravenous; Gunshot woun to chest Yes 2.8 ay binges smoke 7 44 F Every 2 weeks Nasal Methamphetamine Yes M hits per ay Intravenous Gunshot woun to chest Yes M 4 1 Every 2 weeks Intravenous Methamphetamine Yes F lines per Nasal Cariovascular isease plus Yes 9.2 ay methamphetamine toxicity 11 2 M Daily Nasal; oral Methamphetamine Yes 19.8 Yes M 7 18 Daily, limite only by funs Nasal; oral; smoke Cariovascular isease plus methamphetamine toxicity M 11 >8 Daily Smoke Acute aortic issection Yes M 22 >1 Unknown Unknown Methamphetamine Yes F Every few ays Oral; Methamphetamine Yes 7.1 intravenous F 12 >1 Unknown Unknown Methamphetamine Yes 1.8 a Information on subjects 1 12 has been reporte previously by Wilson et al. (9). b Measure in nanomoles (methamphetamine plus amphetamine) per gram of tissue (occipital cortex). centration use maximally stimulate basal aenylyl cyclase activity (19). Gpp(NH)p stimulation was expresse as percentage increase above the basal aenylyl cyclase activity. The titration curves were fitte to the hyperbolic equation to calculate maximal stimulation an EC5 (the concentration proucing 5% of the maximal stimulation), using Origin 5. (OriginLab Corporation, Northampton, Mass.). Data Analysis Statistical analyses were carrie out with two-taile Stuent s t test for inepenent samples an one-way or two-way analyses of covariance (ANCOVA) with age an postmortem interval as the covariates followe by post hoc Tukey honestly significant ifference tests. Pearson prouct-moment correlation or Spearman rank-orer correlation analyses were use to examine the relationships inicate in the text. Results No significant correlations (Pearson) were foun in either group between subject age an levels of any of the outcome measures. There were weak, negative correlations between postmortem interval an basal aenylyl cyclase activity in the absence or presence of GTP (comparison subjects: r=.35 to.1, f=19, p>.11; methamphetamine users: r=.45 to.15, f=14, p>.8) with that in frontal an temporal cortices of the comparison subjects being statistically significant (r=.58 to.45, f=19, p value range=.5 to.4). For the comparison subjects (f=19), no significant correlations were foun between postmortem interval an maximal opamine stimulation in any of the examine brain areas (nucleus accumbens: r=.42, p=.6; cauate: r=.33, p=.14; putamen: r=.15, p=.51; frontal cortex: r=.29, p=.2; temporal cortex: r=.23, p=.31). No significant correlations were seen in the methamphetamine users (f=14) either between postmortem interval an maximal opamine stimulation in any of the examine brain areas (nucleus accumbens: r=.15, p=.57; cauate: r=.2, p=.47; putamen: r=.3, p=.92; frontal cortex: r=.26, p=.33; temporal cortex: r=.2, p=.46). A negative correlation was observe between postmortem interval an maximal Gpp(NH)p stimulation in all of the brain areas (comparison subjects: r=.59 to.36, f=19, p value range=.4 to.11; methamphetamine users: r=.51 to.33, f=14, p value range=.4 to.21), with significant correlations seen in the cauate (r=.59, f=19, p<.5) an frontal cortex (r=.54, f=19, p<.2) of the comparison subjects an in the nucleus accumbens of the methamphetamine users (r=.51, f=14, p<.5). As shown in Table 2, basal activity of aenylyl cyclase (in the presence of 1 µm GTP) was normal in all examine brain regions of the methamphetamine users. Dopamine titration (.4 4 µm) in frontal cortex an Am J Psychiatry 16:5, May 23

4 TONG, ROSS, SCHMUNK, ET AL. TABLE 2. Aenylyl Cyclase Activity Stimulation by Dopamine an Guanylyl-Imioiphosphate [Gpp(NH)p] in Postmortem Brain Tissue of Methamphetamine Users (N=16) an Comparison Subjects (N=21) Dopamine Stimulation Gpp(NH)p Stimulation Brain Region an Basal (plus GTP) (pmol/min per mg) EC5 a (µm) Maximal Stimulation (%) Basal (pmol/ min per mg) EC5 a (µm) Maximal Stimulation (%) Subject Group Mean SD Mean SD Mean SD Mean SD Mean SD Mean SD Nucleus accumbens Comparison Methamphetamine b Cauate Comparison Methamphetamine b Putamen Comparison Methamphetamine b Frontal cortex Comparison Methamphetamine Temporal cortex c Comparison Methamphetamine a The concentration proucing 5% of the maximal stimulation. b Significantly lower than maximal stimulation of comparison subjects (F>12, f=1, 33, p<.2). c Data available only for basal an maximal opamine stimulation. striatum showe the expecte ose epenence an was highly significant (F>2, f=7, 245, p<.1) (Figure 1). No significant ifferences were observe in EC5 between the comparison subjects an the methamphetamine users (Table 2). However, the magnitue of maximal opamine stimulation (calculate from the titration curves) in nucleus accumbens, cauate, an putamen of the methamphetamine users was significantly lower than those of the comparison subjects by 25% 3%, with nonsignificantly lower stimulation seen in the frontal cortex (17% ecrease) an temporal cortex (2% ecrease, calculate from a single saturating concentration of opamine at 1 µm). As shown in Figure 2, although there was extensive overlap between the iniviual comparison an methamphetamine values for maximal opamine stimulation, most of the methamphetamine user values in the striatum fell lower than the mean level for the comparison subjects. Gpp(NH)p titration ( M) was carrie out in the nucleus accumbens, cauate, putamen, an frontal cortex. As escribe in Table 2, no significant ifference was foun between the methamphetamine users an comparison subjects in basal aenylyl cyclase activity, EC5, an the magnitue of Gpp(NH)p stimulation in any of the brain regions. No significant correlations (Spearman rank) were foun between the extent of maximal regional opamine stimulation reporte here an the regional protein concentrations of the opamine D 1 receptor in the 12 examine methamphetamine users in which both measurements were conucte (17), or between maximal opamine stimulation an brain (occipital cortex) levels of methamphetamine plus its metabolite amphetamine in the entire group of 16 methamphetamine users (18) (ata not shown). In aition, there was no significant correlation (Pearson) between opamine stimulation an uration of rug use in those cases (N=11) for which accurate uration information was available (ata not shown). Discussion The major fining of our stuy is that striatal opamine D 1 -stimulate aenylyl cyclase activity is ecrease in human chronic methamphetamine users. We attempte to aress, as much as possible, potential confouning issues associate with postmortem investigations of rug users. Thus, we obtaine forensic evience in bloo, autopsie brain, an, for most of the users, hair, proving that the subjects of our stuy use methamphetamine both recently (rug positive in bloo an brain) an chronically (sequential hair segments). Although the results of rug analyses an retrospective structure interviews suggeste that the rug users use only methamphetamine, it is quite possible that the subjects might previously have use other rugs of abuse or might even have ha a neurochemical efect before rug taking that coul have affecte the biochemical outcome measures. Issues surrouning ifferences in age, postmortem time, an agonal status (suen versus slow eath) were aresse by matching the comparison subjects an methamphetamine users with respect to these variables. We have also previously reporte that the extent of maximal opamine stimulation of aenylyl cyclase activity in biopsie versus autopsie human brain is similar (19). The absence of significant correlation between postmortem interval an opamine stimulation was consistent with the reports on rats (22 24). Our fining of ecrease striatal opamine stimulation of aenylyl cyclase in brain tissue of human methamphetamine users is consistent with investigations reporting be- Am J Psychiatry 16:5, May

5 DOPAMINE RECEPTORS AND METHAMPHETAMINE FIGURE 1. Dopamine Stimulation of Aenylyl Cyclase Activity in Postmortem Brain Tissue of Methamphetamine Users an Comparison Subjects Dopamine Stimulation (%) Nucleus accumbens a Comparison subjects (N=21) Methamphetamine users (N=16) Cauate b Frontal cortex Dopamine (µm) [log scale] a Significant ifference between groups (F=12.5, f=1, 33, p<.2). b Significant ifference between groups (F=18., f=1, 33, p=.2). c Significant ifference between groups (F=6.6, f=1, 33, p<.2). Significant post hoc ifference between groups (p<.4, Tukey honestly significant ifference test). Putamen c low-normal opamine D 1 stimulation of striatal aenylyl cyclase following either acute aministration (25 27) or repeate exposure plus rug challenge (28, 29) of amphetamine to roents. The animal ata suggest that impaire opaminergic stimulation of aenylyl cyclase in humans was not a preexisting abnormality but rather a consequence of exposure to methamphetamine. However, repeate amphetamine aministration to roents can also inuce enhance opamine D 1 receptor-meiate inhibition of nucleus accumbens neurons (3, 31). Although the relevance of these experimental animal stuies employing ifferent treatment paraigms an outcome measures to human rug users is unknown (see reference 32 for review), this suggests that some opamine D 1 receptor-meiate functional changes might be ifferentially affecte by methamphetamine. The cause of the ecrease opamine stimulation of aenylyl cyclase in our human investigation is not known but in principle coul be explaine by ecrease levels of or coupling between the opamine receptor, G protein, an aenylyl cyclase. Our previous observations that striatal levels of opamine D 1 receptor protein are either normal (cauate, putamen) or elevate (nucleus accumbens) in methamphetamine users (17), together with our finings that levels of the stimulatory striatal G protein G olf (33) an basal aenylyl cyclase activity (present investigation) are normal, suggest that ecrease opaminergic stimulation of aenylyl cyclase is unlikely to be explaine by low concentration (e.g., ue to rug toxicity) of these components of the opamine D 1 receptor system. Similarly, the emonstration in methamphetamine-expose roents (25 29) an humans of normal aenylyl cyclase activation by activators (GTP or Gpp[NH]p), which irectly stimulate aenylyl cyclase via G s /G olf, suggests that G protein an aenylyl cyclase coupling is preserve an inicates specificity of the opamine-relate isturbance. However, the possibility cannot be exclue that G protein an aenylyl cyclase coupling associate with opamine receptor activity might have represente only a minor part of total G protein an aenylyl cyclase coupling assesse uner the conitions of our assay. We suggest that esensitization of opamine-stimulate aenylyl cyclase in the methamphetamine users coul be explaine by impaire coupling between the opamine D 1 receptor an the stimulatory G protein (27). Since all of the methamphetamine users use the rug recently as well as chronically (for at least 1 year), receptor esensitization might have occurre consequent to either acute or chronic rug exposure. Although the relative biological importance of the ifferent opamine D 1 -linke effectors (e.g., aenylyl cyclase, phospholipase C [34]) is not known, it is reasonable to expect that much, if not most, of opamine D 1 receptor activity is meiate by G protein activation of aenylyl cyclase (12). Given that methamphetamine causes release of opamine from striatal nerve enings, ecrease opa- 9 Am J Psychiatry 16:5, May 23

6 TONG, ROSS, SCHMUNK, ET AL. FIGURE 2. Maximal Extent of Dopamine Stimulation of Aenylyl Cyclase Activity in Postmortem Brain Tissue of Methamphetamine Users an Comparison Subjects Maximal Dopamine Stimulation (%) Comparison subjects (N=21) Methamphetamine users (N=16) Cauate a Putamen b a Significant ifference between groups (F=22.3, f=1, 33, p<.1). b Significant ifference between groups (F=13.4, f=1, 33, p=.1). c Significant ifference between groups (F=12.3, f=1, 33, p=.1). Nucleus Accumbens c Frontal Cortex Temporal Cortex mine-stimulate aenylyl cyclase activity in methamphetamine users can reasonably be consiere as esensitization (i.e., biochemical tolerance) ue to excessive opaminergic stimulation of the opamine D 1 receptor. However, as the literature is limite on the nature of the involvement between opamine D 1 receptor function an rug-taking behavior in the human, the biological significance of impaire opamine D 1 receptor function in methamphetamine users is uncertain. In this regar, in view of the preliminary evience that the opamine D 1 receptor might meiate part of the euphoric effects of opaminergic psychostimulants in humans, ecrease opamine D 1 receptor function in a limbic brain area (nucleus accumbens) suggests that some D 1 receptor-relate tolerance to the euphoric effects of methamphetamine might occur in human users following repeate rug exposure. It is also possible that subnormal opamine D 1 receptor-aenylyl cyclase activity in this brain area, in aition to the even more severe reuction in tissue stores of striatal opamine (9, 1), might explain part of the ysphoria associate with withrawal from methamphetamine. Finally, the preliminary report escribing ecrease craving for the psychostimulant cocaine in human users following aministration of a selective D 1 agonist (14) also suggests the much more speculative possibility that ecrease opamine D 1 - stimulate aenylyl cyclase activity in limbic brain coul explain, in part, compulsive rug craving, which can occur in some chronic methamphetamine users (1). Previously, we reporte that concentrations of the opamine D 1 receptor were selectively increase in the postmortem nucleus accumbens subivision of the striatum of 12 of the 16 methamphetamine users examine in the present stuy (17). We now fin, however, that espite the increase receptor levels, a functional, an probably more biologically relevant, inex of opamine D 1 receptor activity is below normal in the nucleus accumbens as well as in two other subivisions in which opamine D 1 receptor number was normal. This iscrepancy between receptor number an function suggests that caution shoul be employe in preicting functional changes in receptor activity from ifferences in receptor an G protein concentration. In this regar, the previous reports of changes in several components of the opamine D 2 receptor system in brain tissue of methamphetamine users, e.g., a tren for ecrease in D 2 receptor protein levels (17, 35) an ecrease inhibitory G protein (33), nee to be extene to inclue assessment of whether these changes actually affect D 2 receptor-meiate activity. Unfortunately, however, because of high intersubject variability in our postmortem brain stuy, we were unable to evelop a vali proceure that coul assay opamine D 2 receptor-inhibite aenylyl cyclase activity in autopsie brain homogenates (Tong an Kish, unpublishe observations). We suspect that changes in activity of ifferent opamine (e.g., D 1, D 2 ) an nonopamine receptor systems unerlie some of the behavioral effects of psychostimulant rugs. Our ata provie the first functional ata in human brain suggesting that the opamine D 1 receptor might be one of the systems involve in meiating as yet unetermine aspects of methamphetamine-inuce be- Am J Psychiatry 16:5, May

7 DOPAMINE RECEPTORS AND METHAMPHETAMINE havior. Clinical stuies of pharmacological agents moifying opamine D 1 receptor function in methamphetamine users will establish the biological significance of our neurochemical finings. Receive Feb. 11, 22; revisions receive Aug. 6 an Oct. 29, 22; accepte Nov. 5, 22. From the Human Neurochemical Pathology Laboratory an the Movement Disorer Research Laboratory, Center for Aiction an Mental Health; the Highlan Psychiatric Research Founation, Inverness, Scotlan; the Office of the Santa Clara County Meical Examiner-Coroner, San Jose, Calif.; the Arkansas State Crime Laboratory, Little Rock; the Division of Forensic Toxicology, Arme Forces Institute of Pathology, Washington, D.C.; the Division of Neuropathology, Lonon Health Science Center, University of Western Ontario, Lonon, Ontario; the Spokane County Meical Examiner s Office, Spokane, Wash.; an the Clark County Office of the Meical Examiner, Vancouver, Wash. 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