Brain Research, 543 (1991) 227-235 © 1991 Elsevier Science Publishers B.V. (Biomedical Division) 0006-8993/91/$03.50 A DONIS 000689939116380C

227

BRES 16380

Supersensitivity to the reinforcing effects of cocaine following 6-hydroxydopamine lesions to the medial prefrontal cortex in rats Susan Schenk, Brian A. Horger, Rachel Peltier and Keith Shelton Department of Psychology, Texas A and M University, College Station, TX 77843 (U.S.A.) (Accepted 18 September 1990) Key words: Cocaine; Self-administration; Medial prefrontal cortex; Drug abuse; 6-Hydroxydopamine; Dopamine; Supersensitivity

The effects of neurotoxic lesions to the medial prefrontal cortex on both the acquisition and maintenance of intravenous cocaine self-administration were examined. In one experiment, acquisition of intravenous cocaine self-administration (0.25, 0.5 or 1.0 mg/kg/infusion) was measured in separate groups of rats 14 days following either a sham or 6-hydroxydopamine lesion to the medial prefrontal cortex. For sham rats, the 1.0 and 0.5 mg/kg dose supported reliable self-administration as indicated by discriminative responding. These rats reliably chose a lever that resulted in the delivery of these doses of cocaine over an inactive lever. Reinforced response rates were reduced when 0.25 mg/kg was the available dose and there was a loss of discriminative responding for some of the rats suggesting that it was close to threshold for self-administration. For rats that sustained a 70% depletion of dopamine in the medial prefrontal cortex, the dose-response curve was an inverse function across the entire dose range tested. In contrast to the data from the control rats, lesioned rats had a high rate of reinforced responses and demonstrated good discrimination for all doses including 0.25 mg/kg/infusion, suggesting a supersensitive response to the initial reward effect of cocaine. Another group of rats was first screened for reliable cocaine self-administration (0.5 mg/kg/infusion) and then subjected to either the prefrontal cortical 6-hydroxydopamine or sham lesion. Dose-response curves for cocaine self-administration were compared 14 days following the infusions. The lesioned rats responded reliably for low doses of cocaine that were unable to maintain responding in sham rats. These data support the hypothesis that the medial prefrontal cortex plays an important role in cocaine self-administration.

INTRODUCTION T h e self-administration of cocaine has been linked to the drug's ability to enhance central dopaminergic transmission. In particular, cocaine has indirect agonistic effects through its ability to block the reuptake of d o p a m i n e . This p r o p e r t y has been suggested as the basis for cocaine r e w a r d and m a n y investigators have further suggested that the mesolimbic d o p a m i n e system is the critical one for the self-administration of cocaine 636'21,23. Recently, the notion that the mesocortical d o p a m i n e system plays an i m p o r t a n t role in the self-administration of cocaine has been advanced. Through the usage of intracranial self-administration techniques, G o e d e r s and Smith 8 first r e p o r t e d that cocaine had rewarding properties when directly infused into tissue in the medial prefrontal cortex. Follow-up studies from this l a b o r a t o r y s u p p o r t e d this initial finding and further characterized the rewarding p r o p e r t i e s of cocaine in this neural region to be due to activity in d o p a m i n e systems 93°. If this d o p a m i n e system is critical for the expression of cocaine's rewarding properties, then one would expect that much of the variability in cocaine intake could be explained by

levels of d o p a m i n e or d o p a m i n e activity in the medial prefrontal cortex. H o w e v e r , a neurotoxic lesion to the d o p a m i n e terminals in the medial prefrontal cortex failed to alter intravenous self-administration of cocaine in experienced rats 12. These seemingly discrepant findings can be reconciled in at least 3 ways. First, the two laboratories examined two different aspects of cocaine self-administration. The intracranial technique studied the role of the prefrontal cortical d o p a m i n e system in the acquisition of cocaine self-administration whereas the lesion technique examined the role of this system in the m a i n t e n a n c e of cocaine self-administration. This raises the possibility that cocaine r e w a r d is c o m p r i s e d of an initial and a later phase which m a y be d e p e n d e n t on the activation o f different dopaminergic systems. Second, the lesion study employed only a single dose of cocaine ( a b o u t 0.61 mg/ kg/infusion) which m a y have p r e c l u d e d the ability to assess potential changes in the d o s e - r e s p o n s e curve for cocaine self-administration. Third, there may be something unique about the intracranial route of administration of cocaine when c o m p a r e d to systemic routes of administration.

Correspondence: S. Schenk, Department of Psychology, Texas A and M University, College Station, TX 77843, U.S.A.

228 T h i s p r e s e n t s t u d y is a n i n v e s t i g a t i o n of t h e s e possibilities. I n o n e g r o u p o f rats, t h e ability for 6 - h y d r o x y dopamine

(6-OHDA)

lesions to the medial prefrontal

c o r t e x to a l t e r t h e a c q u i s i t i o n o f i n t r a v e n o u s c o c a i n e s e l f - a d m i n i s t r a t i o n w a s e x a m i n e d . I n a s e c o n d g r o u p of r a t s t h e effect o f t h e s a m e t y p e o f l e s i o n o n m a i n t e n a n c e of c o c a i n e s e l f - a d m i n i s t r a t i o n b y e x p e r i e n c e d r a t s was assessed. In both experiments, dose-response

relations

were determined.

MATERIALS AND METHODS

Expt. 1. Effect of 6-OHDA infusions on the acquisition of intravenous cocaine self-administration Subjects. Subjects were adult male Sprague-Dawley rats (Harlan, TX) weighing approximately 400 g at the time of surgery. They were housed singly in a temperature-controlled colony (22 °C) according to guidelines set forth by NIH. Food and water were available ad libitum. Surgery. The rats were anesthetized with separate injections of sodium pentobarbitol (20 mg/kg, i.p.) and ketamine (60 mg/kg, s.c.). They were then prepared with indwelling jugular cannulae and chronic cannulae (22 gauge) implanted bilaterally in the medial prefrontal cortex (co-ordinates with the upper incisor bar set at -3.9 were: 3.3 mm anterior to bregma; 0.7 lateral; 3.2 ventral to dural. In these rats, a neurotoxic lesion to the medial prefrontal cortex was performed at this time of surgery. Thirty min before an infusion of 6-OHDA HBr (Aldrich Chemical Co.), the rats received an intraperitoneal injection of pargyline (50 mg/kg, Sigma Chemical Co.). Infusions were then delivered via 28 gauge inner cannulae which extended 0.5 mm beyond the tip of the outer cannula. The infusion was controlled via a pump (Harvard Apparatus model PA 22). A 10 /~1 Hamilton syringe was connected to each of the inner cannulae via microbore tubing (Fisher Scientific). The flow of solution was confirmed by observing the movement of an air bubble placed in the microbore tubing. Bilateral infusions of 2 gl per side (4 gg/#l, measured as the base) were made at a rate of 0.4 gi/min. Control rats received infusions of the 0.3% ascorbic acid vehicle. The inner cannula was left in place for 5 min following the infusion. Each day following the surgery, the patency of the catheters was checked by infusing a 0.05 ml solution of saline, heparin (1.25 unitsdml) and penicillin G sodium (250,000 units/ml). Fourteen days following the lesion, behavioral testing began. Apparatus. The self-administration laboratory was equipped with 16 operant chambers (Med Associates). Each box had two levers and two stimulus lights above the levers. They were enclosed in sound attenuating chambers with fans for ventilation. Suspended above each box, was a swivel3 allowing free movement of the rat in the operant box. Depression of one of the levers, designated the 'active' lever, resulted in a 12 s infusion of 0.1 ml of drug through Razel pumps (model A, equipped with 1 RPM motors and 20 ml syringes). Depression of the other level, designated 'inactive', had no consequence. The stimulus light above the active lever was activated during the infusion. The system was interfaced with two microcomputers (Med Associates). Control of drug delivery and data acquisition was obtained through the OPN software package (modified from refs. 7 and 19). Procedure. Some rats (13 lesion and 15 control) were sacrificed 14 days following the lesion or sham lesion procedure (see section on Biochemical assays) in order to assess the neurochemicai effects of 6-OHDA in cocaine-naive rats. Others were tested for the spontaneous acquisition of lever pressing for cocaine infusions. These rats were placed in the operant boxes on a limited access schedule of 2 h daily, without having received any prior lever press training. On

each day, the first administration of cocaine was experimenter delivered and served as a 'priming' infusion. Thereafter, depression of the active lever resulted in the delivery of an infusion of 1.0, 0.5 or 0.25 mg/kg/infusion cocaine HC1 (Sigma Chemical Co.) dissolved in physiological saline.

Expt. 2. Effect of 6-OHDA infusions on the maintenance of intravenous cocaine self-administration Subjects. Subjects were 36 male Sprague-Dawley rats weighing approximately 350 g at the time of surgery. They were housed individually in standard plastic hanging rodent cages in a temperature controlled animal colony (22 °C). Food and water were freely available. Surgery. The rats were prepared with jugular cannutae and bilateral medial prefrontal cortical cannulae as in Expt. 1 except that the intracranial cannulae were fitted with internal dummy cannulae to maintain their patency. The 6-OHDA or sham lesion was performed at a later date (see phase 2 below). The rats were permitted at least 6 days to recover from the surgical manipulation. On each of these days the patency of the catheter was confirmed by infusing a 0.1 ml solution of saline, streptokinase (700 units/ml, Sigma Chemical Co., St. Louis, MO), heparin (30 units/ml) and penicillin G sodium (125,000 units/roll. Procedure. The experiment was divided into 3 phases. The first, an acquisition phase served to screen rats for cocaine selfadministration. The second phase involved a waiting period of 14 days following either 6-OHDA or sham lesion to the medial prefrontal cortex. The third phase involved retesting of the lesion and sham rats to assess the effect of the lesion.

Phase 1: acquisition of cocaine self-administration Each of these 2 h test days began with an experimenter delivered priming infusion. Thereafter, infusions were earned on an FR1 schedule by depressing one of two levers. Depression of the other 'inactive' lever was without consequence. On each of 7 days, 0.5 mg/kg/infusion was the available dose. There were 24 of the initial 36 rats (67%) that demonstrated a preference for the active lever (greater than 50% active lever responses). This figure is consistent with a recent study that also reported 67% of rats self-administered cocaine 2. Thus the variability in responding to the reinforcing effect of cocaine obtained in our laboratory is consistent with that obtained in another laboratory. These rats were deemed sensitive to cocaine's reinforcing effects and were randomly assigned to the lesion or sham condition.

Phase 2: lesion Rats were lightly anesthetized with ketamine (60 mg/kg, s.c.). Thirty minutes before an infusion of 6-hydroxydopamine HBr (Aldrich Chemical Co.) or its 0.3% ascorbic acid vehicle, the rats received an intraperitoneal injection of pargyline (50 mg/kg, Sigma Chemical Co.). The first 8 rats treated with this combination of drugs died. Although we had administered these treatments without incidence before (Expt. 1), we had never done so in cocaine experienced animals. Thus it is possible that chronic exposure to cocaine sensitized rats to the lethal effects of ketamine and/or pargyline. Due to this unfortunate consequence, the remaining rats were pretreated only with pargyline, without incidence. Infusions were delivered via 28 gauge inner cannulae as above. A waiting period of 14 days insued. During this time, the jugular cannulae were flushed daily with the heparin, streptokinase, penicillin solution. Following this 14 day period, the rats were again tested to determine the effect of the lesion on cocaine selfadministration.

Phase 3: retest of cocaine self-administration On the first 3 days, the available dose was 0.5 mg/kg/infusion. Thereafter, the dose was repeatedly reduced by half every 2 days. On the final test day, saline was substituted for the cocaine solution. Thus on days 1-3, the dose was 0.5 mg/kg, on days 4 and 5 it was 0.25 mg/kg, on days 6 and 7 it was 0.125 mg/kg, on days 8 and 9 it

229 14 days following the sham lesion served to define an

TABLE I

Neurochemical effects of a 6-OHDA or ascorbic acid vehicle (control) infusion into the medial prefrontal cortex Data are presented for rats that either completed the entire testing procedure (cocaine experienced: n = 29 6-OHDA; n = 26 sham) or for rats that were sacrificed prior to any behavioral testing (cocaine naive: n = 13 6-OHDA; n = 15 sham). Mean values are expressed as ng/gwet tissue + S.E.M.

Cocaine experienced DA DOPAC/DA NE Cocaine naive DA DOPAC/DA NE

effective lesion. A 95% confidence interval was placed about the mean medial prefrontal cortical d o p a m i n e content in these rats. The lower limit of this interval was 57 ng/g. A n y 'lesion' rat that d e m o n s t r a t e d higher levels of dopamine than this value was considered not to have been effectively lesioned and was discarded from the study. The resulting sample sizes per dosage group were:

6-OHDA

Control

30.14 _+3.1 0.58 + 0.09 156.99 + 37.41

100.3 + 4.4 0.29 + 0.02 470.42+_24.84

infusion: lesion -- 11, control = 8. For all neurochemical measures, cocaine exposed and

34.46 + 9.1 0.34 + 0.05 111.85 + 50.44

98.56 -+ 5.0 0.18 + 0.02 488.58+ 31.78

depletions averaged about 70% in the lesioned rats.

1.0 mg/kg/infusion; lesion = 8, control = 8; 0.5 mg/ kg/infusion: lesion =

was 0.06 mg/kg and on day 10 it was saline (0 mg/kg). Four jugular cannulae developed clots or leaks during phase 3. The data from these rats were excluded from further analysis. The remaining 12 subjects (5 sham and 7 lesion) completed the entire 31 day procedure.

RESULTS

Expt. 1. Effect o f the lesion on acquisition o f cocaine self-administration Table I shows the effects of injections of 6 - O H D A into the medial prefrontal cortex on tissue levels of dopamine, D O P A C and norepinephrine. Data from cocaine exposed and n o n exposed rats are presented. The levels of dopamine in the PFC of the 15 rats that were sacrificed

10; 0.25 mg/kg/

cocaine naive rats were markedly similar. D o p a m i n e There was a concomitant increase in the turnover of dopamine as indicated by the significant increase in D O P A C : d o p a m i n e ratios. In addition, the 6 - O H D A lesioned rats had significantly decreased n o r e p i n e p h r i n e levels. Fig. 1 shows the n u m b e r of reinforced (active) and

Biochemical assays One day following the last self-administration test the rats from both experiments were decapitated and the brain rapidly removed. For the rats from the first experiment, only a sample of the medial prefrontal cortex was obtained. For the rats from the second experiment, both a sample of the medial prefrontal cortex and a sample of the nucleus accumbens were dissected on an ice cold plate. The samples were weighed and kept at -90 *C in 1.5 ml microcentrifuge tubes until high performance liquid chromatography (HPLC) analyses were conducted. Tissue samples were prepared on ice by first adding 0.16 N perchioric acid (PCA) containing 100 ng/ml of the internal standard, dihydroxybenzylamine (DHBA, Sigma Chemical Co.). Samples were then sonicated and centrifuged for 2 rain at 14,000 rpm. They were kept refrigerated and 20/A supernatant of each sample was assayed for dopamine, dihydroxyphenylacetic acid (DOPAC) and norepinephrine content using a BAS 400 model HPLC system with LC-4B amperometric detector. Samples from the second experiment were also assayed for serotonin and 5-hydroxyindoleaceticacid content. The monoamines were separated with a reverse phase IIODS (100 × 3.2 ram, 3/*m) column heated at 36 °C. The mobile phase consisted of methanol buffer prepared by mixing 0.0186 g disodium phosphate, 2.8 g citric acid, 200/,1 sodium octyl sulfate (100 mg/ml), 0.946 g EDTA and 970 mi double distilled water. This preparation was filtered and degassed, after which 30 ml HPLC grade methanol was added. The electrode voltage was set at -0.7 V and the indifferent electrode was Ag/AgCI. All samples were run in duplicate.

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230 TABLE II

200 Pre-Lesion I

A N O V A summary table for analysis conducted on reinforced and non-reinforced response rates for each condition t,-

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non-reinforced (inactive) lever presses during the 2 h daily self-administration sessions. The left panel presents the data from the control rats and the right panel presents the data from the lesioned rats. Data are presented for the different available doses of cocaine (1.0, 0.5 or 0.25 mg/kg/infusion). On day 1, the inactive lever responses tend to be high and there is no evidence of discriminative responding. Over days, inactive lever responding extinguishes and responding is confined almost exclusively to the active lever. Thus, it appears that the rats learned to ignore irrelevant stimuli (the inactive lever) in their environment. To analyze whether the rats had reliably learned to discriminate the active and inactive levers, we performed 6 individual ANOVAs comparing the number of active and inactive lever presses from each panel in Fig. 1. The results are summarized in Table 1I. For the control rats, the difference between the number of reinforced and non-reinforced responses was significant for the 1.0 and 0.5 mg/kg/infusion groups. The

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Fig. 3. Reinforced responses for various doses of cocaine in 6-OHDA and sham lesioned rats. The data were obtained 14 days following the infusion. At the extreme right, the average number of reinforced responses on the last two prelesion sessions are presented. The bars represent the average number of responses for the lesioned and control rats. Symbols represent the individual rat data. Responding for the lower dose (0.06 mg/kg/infusion) was higher for the 6-OHDA lesioned rats. The difference in responding for 0.125 and 0.25 mg/kg/infusion approached significance.

difference failed to reach significance for the 0.25 mg/kg/infusion group. For the lesioned rats, the difference in response rates was significant for all 3 doses tested. Additional ANOVAs were conducted to compare number of reinforced lever presses for control and lesioned rats tested with each dose of cocaine. Rats tested with the highest two doses (1.0 and 0.5 mg/kg/infusion) did not differ in responding as a function of lesion condition. In contrast, marked differences were observed at the 0.25 mg/kg/infusion dose. The lesioned rats

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231 exhibited more responding during the later days of the 8 day acquisition period• The control rats had a low and relatively invariant number of responses throughout the 8 days. A 2-way A N O V A (Lesion condition × Days, with repeated measures on the days factor) yielded a significant main effect of lesion condition (Fl,]7 = 4•281, P = 0.05)• The interaction between lesion condition and days approached significance (F7,n9 = 1•625, P = 0•13)• Analyses of simple main effects revealed that the lesioned rats had a higher number of responses on day 5 (F1,51 = 5.7, P < 0.01), day 6 (F1,51 = 7.306, P < 0.01), day 7 (F1,51 = 5.091, P < 0.01) and day 8 (Fl,51 = 4.371, P < 0.05).

Expt. 2. Effect of the lesion on maintenance of cocaine self-administration The biochemical analyses revealed that two of the 'lesion' rats that completed the entire testing procedure had not sustained significant depletions of dopamine following the 6 - O H D A infusion. When compared to the sham rats that completed testing, their depletions were only 23 and 17%. The remaining 5 subjects sustained an average of 56.4% dopamine depletion (range 46-65%) in the medial prefrontal cortex. Fig. 2 shows the results of

the biochemical analyses on prefrontal cortical and nucleus accumbens samples from the rats that completed all 3 phases of the experiment. Only the 5 subjects that sustained greater than a 45% depletion are included in the 'lesion' group. As can be seen, there were substantial depletions of prefrontal cortical dopamine, D O P A C and norepinephrine in the lesioned rats. In addition, there was a small decrease in the levels of serotonin and in the DOPAC:dopamine ratio• The lesion was without effect on these neurochemicals in the nucleus accumbens. Fig. 3 shows the number of reinforced responses for lesioned and sham rats 14 days following the lesion to the medial prefrontal cortex. Data are presented for the last test day at each of the doses. For comparison, the average number of reinforced responses on the last two prelesion days are also presented at the extreme fight of the figure. For the prelesion days, individual data for the 5 lesion and 5 sham rats are presented as symbols within the bars that represent the average number of responses• Marked differences in responding for the lower doses of cocaine were found following the lesion• A 2-way A N O V A (Condition x Dose, with repeated measures on the Dose factor) revealed a significant effect of Condition (F1, 8 = 9.641, P = 0.0146), Dose (F4,32 = 6.443, P =

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0.0006) and an interaction between the two variables (F4,32 = 4.271, P = 0.007). Simple main effect analyses revealed that the difference in responding for the 0.06 mg/kg dose was highly significant (F1, 8 = 8.02, P = 0.022). Differences in responding for the 0.125 and 2.5 mg/kg doses approached significance (F1. 8 = 4.389, P = 0.069; F1,8 = 3.986, P = 0.081, respectively). Fig. 4 shows the percentage of total lever responses devoted to the active lever. Again, the data are presented for the last test day for each dose. These data suggest that the increase in responding observed following the lesion was not simply an artifact of an increase in non-specific activity since responding on the inactive lever was not similarly affected. These responses remained low for all doses of cocaine tested, as indicated by the high percentage of active lever responses across all doses of cocaine (greater than 80%). Only when saline was substituted for cocaine did inactive lever responding increase, bringing the percentage of active lever presses to chance levels. Fig. 5 presents scatterpiots of prefrontal cortical dopamine levels and number of reinforced response per session for each cocaine dose for all 12 rats that completed testing. The best fitting linear function is

depicted for each dose. For the lowest doses of cocaine tested (0.06, 0.125 and 0.25 mg/kg/infusion), dopamine levels account for a meaningful percentage of the variability in self-administration (r = -0.57 (0.25 and 0.125 mg/kg), r = -0.65 (0.06 mg/kg), P < 0.05 for each of these doses). This relationship breaks down for the highest (0.5 mg/kg/infusion, r = -0.35) and lowest (0.0 mg/kg/infusion r = -0.18) doses tested, at which the variability in responding is relatively low. Thus at doses which exhibit variability in responding (and by extension, reflect variability in sensitivity to cocaine's effect) whether or not a rat will respond for the drug appears to be inversely related to dopamine levels in the medial prefrontal cortex, which were manipulated by the 6O H D A lesion. Fig. 6 presents scatterplots of medial prefrontal cortical norepinepbrine concentration and reinforced responses for each of the doses of cocaine. The best fitting linear functions for the data are also presented. Although there was substantial variability in norepinephrine concentration, ranging from 15.6 to 547.4 ng/g, this variable did not explain a significant proportion of the behavioral variability. At best, 17% of the variability (r = -0.32) in

233 self-administration of the 0.06 mg/kg/infusion dose was accounted for by norepinephrine levels. DISCUSSION In the first experiment, the effect of a 6-OHDA lesion on acquisition of cocaine self-administration was assessed. These rats received the lesion prior to any training with cocaine. Under these conditions, the majority of sham lesioned rats discriminated between depression of an active lever which delivered an intravenous infusion of 1.0 or 0.5 mg/kg/infusion doses of cocaine and an inactive lever. Thus most of the rats in these two dosage groups were reliably self-administering cocaine. Many of the sham lesioned rats failed to effectively discriminate between the active and inactive levers when 0.25 mg/kg/infusion served as the reinforcer. As such, the variability in number of reinforced responses was high in this group (range of reinforced responses on day 8 was 1-48) and a comparison of reinforced and non-reinforced responding indicated that this dose of cocaine was too low to support reliable responding for the group as a w h o l e , and is thus near the threshold for self-administration. Had additional lower doses of cocaine been tested in this experiment the threshold dose would have more convincingly been established. A different profile was obtained for the lesioned rats. Whereas reinforced responses for the 1.0 and 0.5 mg/ kg/infusion doses were comparable to the rate of the control rats, there was a large and significant difference in reinforced responding for the 0.25 mg/kg/infusion dose. The larger number of responses produced by the lesioned group as a w h o l e was primarily due to the larger number of lesioned rats that responded for this dose of cocaine. The difference in responding for the 0.25 mg/kg dose as a function of lesion condition suggests that the dose-response curve for the lesioned rats was displaced to the left. However, a more convincing test of this hypothesis would require a comparison of the behavior of control and lesioned rats during acquisition of cocaine self-administration with additional low doses (less than 0.25 mg/kg) of cocaine available. In the second experiment, rats sustained a neurotoxic lesion of the medial prefrontal cortex following initial acquisition of cocaine self-administration. In these rats, a wider range of doses was tested. When the available dose of cocaine was reduced below 0.25 mg/kg, the average number of responses per session decreased for the sham lesioned rats, eventually reaching the low rate comparable to when saline is substituted for cocaine. The gradual rather than abrupt decrease in responding with decreases

in dose is a reflection of a gradual decrease in the number of rats that maintain responding for the lower dose of drug. In contrast to the data from the control rats, the number of reinforced responses for lesioned rats continued to increase in a compensatory fashion as the available dose of cocaine was reduced below 0.25 mg/kg. The higher average number of responses for these rats is a reflection of the higher number of rats that responded reliably for these low doses of cocaine. Thus the threshold for the rewarding effects of cocaine was lower for this group than for the control rats. It is of interest that most of the control rats from the second experiment reliably responded for the 0.25 rag/ kg/infusion dose of cocaine that had been subthreshold for many rats in Expt. 1 (see Fig. 1). As a result, the average number of responses for this dose of cocaine was higher than had been observed in the first experiment. One difference that may explain these different results is that rats from the second experiment received substantial exposure to cocaine prior to receiving the 0.25 mg/ kg/infusion dose (7 days prelesion and 3 days postlesion). These exposures may have served to sensitize these rats to subsequent cocaine infusions such that they were more responsive to the lower doses of cocaine. Rats that have had experience with cocaine tend to be more sensitive to subsequent effects of the drug as measured in tests of cocaine induced locomotor activity 15. We have recently reported such a supersensitive response to the rewarding properties of cocaine in rats that had been pretreated with systemic injections of cocaine 11. Thus, it is possible that the control rats from the second experiment responded differently to cocaine due to this prior exposure. When saline was substituted for the cocaine solution, responding dropped off markedly for the lesioned rats, to rates comparable to those of the control rats. In addition, inactive lever responses remained low and were not similarly elevated. Taken together these data argue that the increase in responding was specific, that the animals maintained their ability to discriminate and that the effects of the 6-hydroxydopamine infusion into this dopamine terminal area had induced a supersensitivity to cocaine's reward effect. In both experiments, the lesion produced marked depletions of both dopamine and norepinephrine in the medial prefrontal cortex. We failed to protect the noradrenergic terminals from the neurotoxin since, in our preliminary experiments, pretreatment with the uptake inhibitor, desmethylimipramine, caused many of our rats to become extremely ill. However, a role of norepinephrine in mediating the rewarding effects of cocaine is unlikely since systemic administration of specific noradrenergic receptor blockers were ineffective in altering

234 intravenous cocaine self-administration 6. Also, local application of antagonists failed to effect the intracranial self-administration of cocaine directly into the medial prefrontal cortex 9. Further, the variability in self-administration of any dose of cocaine tested was not linearly related to prefrontal cortical n o r e p i n e p h r i n e levels (see Fig. 6) although d o p a m i n e levels did explain a significant p r o p o r t i o n of the variability in reinforced response rate for low doses of cocaine (see Fig. 5). Thus, a more likely possibility is that the reduction of d o p a m i n e in this terminal area was at least partly responsible for the differences in sensitivity to cocaine that were observed. Behavioral supersensitivity to d o p a m i n e agonists has previously been o b s e r v e d following subtotal depletions in medial prefrontal cortex 4'17, stratum 2° or in whole brain TM p r o d u c e d by 6 - h y d r o x y d o p a m i n e infusions. The effect has been i n t e r p r e t e d as due to c o m p e n s a t o r y changes that occur in the remaining intact cells. For example, partial lesions p r o d u c e a m a r k e d decrease in high affinity d o p a m i n e uptake and an increase of in vivo d o p a m i n e biosynthesis 1'4'13"22. With more extensive lesions, supersensitivity of postsynaptic mechanisms occurs 5. All of these mechanisms provide an increase in the effectiveness of remaining elements of the d a m a g e d d o p a m i n e system. This increase may m o r e than compensate for the damage done to the system, as indicated by the supersensitive behavioral responses to d o p a m i n e agonists, particularly when greater than 20% of d o p a m i n e terminals still remain intact. This would explain the direction of the change in sensitivity to the reinforcing impact of cocaine p r o d u c e d by our lesion of the medial prefrontal cortex. Further, the REFERENCES 1 Altar, C.A., Marien, M.R. and Marshall, J.E, Time course of adaptations in dopamine biosynthesis, metabolism, and release following nigrostriatal lesions: implications for behavioral recovery from brain injury, J. Neurochem., 48 (1987) 390-399. 2 Bozarth, M.A., Murray, A. and Wise, R.A., Influence of housing conditions on the acquisition of heroin and cocaine self-administration in rats, Pharmacol. Biochem. Behav., 33 (1989) 903-907. 3 Brown, Z.W., Amit, Z. and Weeks, J.R., Simple flow-thru swivel for infusions into unrestrained animals, Pharmacol. Biochem. Behav., 5 (1976) 363-365. 4 Carter, C.J. and Pycock, C.J., Behavioral and biochemical effects of dopamine and noradrenaline depletion within the medial prefrontal cortex of the rat, Brain Research, 192 (1980) 163-176. 5 Creese, I., Burt, D.R. and Snyder, S.H., Dopamine receptor binding enhancement accompanies lesion-induced behavioral supersensitivity, Science, 197 (1977) 596-598. 6 DeWit, H. and Wise, R.A., Blockade of cocaine reinforcement in rats with the dopamine receptor blocker pimozide, but not with the noradrenergic blockers phentolamine or phenoxybenzamine, Can. J. Psychol., 31 (1977) 195-203. 7 Emmett-Ogelsby, M.W., Spencer, D.G. Jr. and Arnoult, D.E., A TRS-80 based system for the control of behavioral experi-

data support the idea that similar c o m p e n s a t o r y mechanisms that function in o t h e r central d o p a m i n e systems also function in the mesocorticai d o p a m i n e system. These data may also provide an indication as to why a previous study failed to observe an effect of m o r e extensive lesions to the medial prefrontal cortex ~2. The dose of cocaine that was used in that study (about 0.61 mg/kg/infusion) fell b e t w e e n the two highest doses tested in the present study. Since we found no effect of the lesion on the self-administration of these two doses of cocaine, it is possible that the choice of a single drug dose in the Martin-Iverson et al. study 12 study obscured any possible change in the d o s e - r e s p o n s e curve. H a d several doses been tested, an effect m a y have been observed. The present data d e m o n s t r a t e that the medial prefrontal cortical d o p a m i n e system plays an i m p o r t a n t role in both the acquisition and m a i n t e n a n c e of intravenous cocaine self-administration. A s a result, d a t a o b t a i n e d following 6 - h y d r o x y d o p a m i n e lesions to the mesolimbic d o p a m i n e terminal a r e a in the nucleus accumbens must be i n t e r p r e t e d with caution, since these lesions also produce m a j o r deficits in d o p a m i n e content in the medial prefrontal cortex 23. Acknowledgements. This project was supported by a NIDA research grant to S.S. (DA 05548). The authors gratefully acknowledge the help and unending support of Dr. T.H. Champney in obtaining the neurochemical data. We also thank Dr. P.J. Wellman for critical evaluation of earlier drafts of the manuscript. The valuable information concerning effective concentrations of heparin and the addition of streptokinase to maintain jugular cannula patency contributed greatly to minimizing subject attrition in Expt. 2. We are grateful to Dr. M. Emmett-Ogelsby for sharing this information.

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235 administration in the rat, Nature, 260 (1976) 731-732. 16 Roberts, D.C.S., Corcoran, M.E. and Fibiger, H.C., On the role of ascending catecholamine systems in intravenous selfadministration of cocaine, Pharmacol. Biochem. Behav., 6 (1977) 615-620. 17 Robinson, R.G. and Stitt, T.G., Intracortical 6-hydroxydopamine induces an asymmetric behavioral response in the rat, Brain Research, 213 (1981) 387-395. 18 Schoenfeld, R.I. and Uretsky, N.J., Enhancement by 6-hydroxydopamine of the effects of DOPA upon the motor activity of rats, J. Pharmacol. Exp. Ther., 186 (1973)616-624. 19 Spencer, D.G., Jr. and Emmett-Ogelsby, M.W., Parallel processing strategies in the application of microcomputers to the behavioral laboratory, Behav. Res. Meth. Instr, 17 (1985)

294-300. 20 Thornburg, J.E. and Moore, K.E., Supersensitivity to dopamine agonists following unilateral, 6-hydroxydopamine-induced striatal lesions in mice, J. Pharmacol. Exp. Ther., 192 (1975) 42-49. 21 Wise, R.A. and Bozarth, M.A., A psychomotor stimulant theory of addiction, Psych. Rev., 94 (1987) 469-492. 22 Zigmond, M.J., Acheson, A.L., Staehowiak, M.K. and Strickerm, E.M., Neurochemieal compensation after nigrostriatal bundle injury in an animal model of preclinical Parkinsonism, Arch. Neurol., 41 (1984) 856-861. 23 Zito, K.A., Vickers, G. and Roberts, D.C.S., Disruption of cocaine and heroin self-administration following kainic acid lesions of the nucleus accumbens, Pharmacol. Biochem. Behav., 23 (1985) 1029-1036.

Supersensitivity to the reinforcing effects of cocaine following 6-hydroxydopamine lesions to the medial prefrontal cortex in rats.

The effects of neurotoxic lesions to the medial prefrontal cortex on both the acquisition and maintenance of intravenous cocaine self-administration w...
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