Reduced behavioral responses to intranigral muscimol following chronic ethanol

Physzology& Behavior,Vol. 46, pp 473-477 ©Pergamon Press plc, 1989 Pnnted m the U S A

0031-9384/89 $3 00 + .00

Reduced Behavioral Responses to Intranigral Muscimol Following Chronic Ethanol I L A R R Y P. G O N Z A L E Z 2 A N D J A N E T F. C Z A C H U R A

University o f Oklahoma Health Sciences Center Department o f Psychiatry and Behavioral Sciences, Oklahoma City, O K 73190 R e c e i v e d 27 O c t o b e r 1988

GONZALEZ, L P. AND J. F. CZACHURA. Reduced behavzoral responses to intramgral musctmolfollowing chronic ethanol. PHYSIOL BEHAV 46(3) 473-477, 1989.--Increased b~ochemical measures of GABA activity are observed after acute administration of ethanol and decreased activity has sometimes been observed after chronic ethanol exposure. Since chronic alterations in neurotransmitter activity may result m changes in receptor function, tt Is possible that changes in GABA-receptive neurons may accompany chromc ethanol treatment In the present study we examined the incidence of musclmol-mduced motor behaviors m ethanol-naive and chromc ethanol-treated animals. Male Sprague-Dawley rats received bilateral cannula implants into substantia nigra pars renculata for subsequent adnunlstratmn of musclmol or saline After recovery from surgery, rats received chronic treatment in ethanol-vapor inhalation chambers for 15 days Animals were then removed from the chambers and examined 10 hours after removal. MUSClmOlresulted m a general increase in motihty in both control and ethanol-treated animals Animals withdrawn from chrome ethanol exposure, however, exhibited slgmficantly less muscimol-stlmulated, repetlnve 9 Hz movements These results suggest that GABA receptwe cells within the substantla nlgra or its VlClnltymay be functionally less responsive to GABAerglc sttmulataon after chronic ethanol adImmstrauon. Withdrawal

Muscimol

Substantaamgra

GABA

Stereotypy

Locomotion

Motility

These chronic effects of ethanol on GABAergic systems are opposite to the effects of acute ethanol administration, and thus st;ggest an adaptation of GABAergic neurons to the chronic presence of ethanol. Behavioral effects of GABA agonists on motor incoordination (3,26) and on spontaneous locomotor activity (36) are, in fact, reduced in animals chronically exposed to ethanol. Treatments that increase GABA activity in animals withdrawn from chronic ethanol treatment have been observed to block handling-induced convulsions in mice (11,12) and audiogenic seizures in rats (10,18). Not all withdrawal symptoms, however, are affected by the administration of GABA agonists, with tremors in ethanol-withdrawn rats and monkeys unaffected by such treatment (10,37). It is thus possible that at least some of the symptoms of ethanol withdrawal may result from an altered response of GABA-receptive neurons. Results from this laboratory (18) suggest that modification of GABAergic activity in the substantia nigra in particular may alter andiogenic seizure susceptibility during ethanol withdrawal. In these studies, intranigral administration of the GABA agonist muscimol was found to cause a significant blockade of audiogenic seizures during ethanol withdrawal. Because of the important effects of acute and chronic ethanol on the function of GABA systems and the implication of GABAergic involvement in the propagation of convulsive seizure activity, the present studies examined the functional responsiveness of nigral neurons to a locally-applied GABA agonist after chronic

GABA (gamma-amino-butyric acid) is one of the principal inhibitory neurotransmitters m the central nervous system (21), and it has been suggested that GABA may play a role in the depressant actions of acute ethanol. Evidence has accumulated over the last few years that ethanol does alter the function of GABAergic systems. Microiontophoretically-applied or systemically-applied ethanol potentiates both GABA-induced inhibition of the singleunit activity of cortical neurons and also the cortical inhibition induced by electrical stimulation of the cortical surface (27,28). Acute ethanol increases GABA binding to a low affinity binding site (39,40), alters steady-state GABA levels (43), and may reduce GABA turnover in the CNS (44). In addition, some of the acute behavioral effects of ethanol are altered by GABAergic drugs. GABA antagonists blocked, and GABA agonists enhanced, ethanol-induced impairment of motor coordination in rats (9, 19, 22), and ethanol effects on sleep, locomotion, and thermal regulation (23). While acute ethanol is reported to increase low-affinity GABA receptor binding, a decreased binding affinity (42,43), and a reduced density of binding sites (40,43) are reported after chronic ethanol treatment. The changes in GABA binding affinity following withdrawal are reported to be correlated with andiogenic seizure susceptibility (39) and with the occurrence of spontaneous convulsions (29), suggesting that the depressed GABA function may be related to these components of the withdrawal syndrome (39).

~Supported in part by Nataonal Insntute on Alcohol Abuse and Alcoholism grant AA006254 to L. P Gonzalez 2Requests for reprints should be addressed to Larry P Gonzalez, University of Oklahoma Health Sciences Center, Research Braiding 306-R, P.O. Box 26901, Oklahoma City, OK 73190-3000

473

474

GONZALEZ AND CZACHURA

HG 1 Frontal section showing typical site of bilateral cannula taps within the substantla mgra pars retaculata

ethanol exposure and withdrawal. Since the intranigral administration of GABA agonists results in alterations in motor behawor and in stereotyped movements (1,33), these studies measured changes in an automated measure of motor behavior as an indication of the function of GABA-receptlve neurons. If the sensitivity of nigral elements to GABA is reduced after withdrawal from chronic ethanol exposure, as suggested by previous reports of neurochemical changes, measurements of stereotyped motor movements following intranigral GABA agonists would also be expected to be reduced. METHOD

Subjects The subjects of these experiments were 32 male, SpragueDawley rats, 60 to 90 days old, and weighing 200 to 250 g Animals were housed in indiwdual cages with free access to food and water, and were maintained for at least seven days under the same conditions of environment, diet, and daily handling before any experimental treatment

Cannula Implantation Animals were prepared with chronic bilateral guide cannulae for subsequent placement of internal lnjectmn cannulae into the substantia nigra pars reticulata (SNR). Cannulae implants were done stereotaxically under sodium pentobarbital anesthesia (40.0 mg/kg, IP, supplemented with chloral hydrate as needed), with 25-gauge guide carmulae placed 5.3 mm posterior to Bregma, 7.0 mm below skull surface, and 2.3 mm lateral to midline, according to the atlas of Paxinos and Watson (30). The chronicallyimplanted cannulae provided for subsequent placement of 32gauge injection cannulae, which protruded one mm beyond the end of the guide, into SN (5.3 mm posterior to Bregma, 8.0 mm below skull surface, and 2.3 mm lateral to midhne). Cannula placements were verified histologically at the completion of this study (see Fig. 1).

Chronic Ethanol Exposure Following a one-week recovery penod after cannula implantation, the animals were divided into two groups of 16 animals per group, including an ethanol-naive control group and a group of chronic-ethanol animals. The chronic-ethanol animals received chronic ethanol exposure in ethanol-vapor inhalation chambers. The chambers are 24-inch Plexiglas cubes subdivided vertically at

12 inches to give two cages (24 × 24 × 12 in.), providing individual housing for four rats. The air flow to each cage was regulated independently. Fresh air was flushed through the chamber continuously at the rate of two hters per minute to provide for the respiratory needs of the animals. To this fresh air flow was added ethanol-saturated vapor (0 to 30 mg/ml) obtained by pumping air through a one-liter aspirator bottle containing 1000 ml of 95% ethanol, at flow rates of 0-700 ml/min. The ethanol flow rate was adjusted to maintain behavioral levels of intoxication between Ataxia-2 to Ataxm-3 as described by Majchrowicz (24). Using this procedure, blood ethanol levels were increased gradually, approaching 350 mg/dl after 15 days of exposure, at which times the animals were removed from the chambers. Control animals received similar handling and housing for the same length of time as did the chronic-ethanol animals, but received no exposure to ethanol. All animals had free access to food and water throughout this penod. Blood ethanol levels were determined periodically during chromc ethanol exposure and following ethanol withdrawal by a head-space gas chromatographic method. In this procedure, 20-ml samples of blood obtained from the dorsal tall vein were added to 1.0 ml of delonized water m a 25-ml flask, with 0.025 mg of propanol added as an internal standard. The flask was lmmedmtely sealed and warmed in a water bath (50°C) for 15 mm. A 1.0-ml sample of the gas volume from the flask was rejected into the port of a Hewlett-Packard 5790-A gas chromatograph equipped with a flame ionization detector. Ethanol levels were quantified by comparison to standards prepared by adding known amounts of ethanol to the propanol solution Blood samples were also obtained from control animals so as to ensure the similarity of handling between controls and chromc-ethanol animals, although ethanol was never detected in the blood of control animals. After 15 days of ethanol-vapor exposure, the ammals were removed from the chambers. Ten hours after removal, motihty measurements were performed as described below

Assessment of Motlh~ Ten hours after the withdrawal of ethanol, quantitative assessment of motor behavior was performed in a Stoelting activity monitor, modified to permit the quantificataon of repetmve behaviors (8,13). The motility monitor consists of a pair of parallel copper plates (20.0 cm × 30.0 cm) connected to a Stoeltmg movement sensor module. The plates are housed in a 40.0 × 40.0 × 40.0 cm Faraday cage to eliminate the influence of external electrical fields. An animal was placed m the center of a radio frequency capacitance field generated between the plates such that the movement of the animal disrupted the field. Based upon this disruption, the mouhty monitor produced an analog signal with a frequency of oscillation equal to the frequency of occurrence of movements within the field of the monitor. This signal was then filtered and amplified with a Grass 7P511 amplifier and recorded on an FM tape recorder for subsequent playback and analysis. Quantification of movement was accomplished by spectral analysis of the frequency components of the amplified analog output of the motility momtor. The resulting amplitude-frequency distribuUon has been shown (7, 8, 13) to accurately depict the occurrence of specific repetitive movements (sniffing, licking, head bobbing, etc.). For the measurement of motility, animals were placed m Plexiglas chambers (19 × 13 × 8.0 cm) that were then positioned within the movement sensor of the motihty monitor. After a 30-minute adaptation period, a 32-second sample of motility was obtained. Ammals then received bilateral intranigral injections of either sahne (0.5 ixl/side) or muscimol (30 ng/side, dissolved in

CHRONIC ETHANOL AND INTRANIGRAL MUSCIMOL

0.5 )xl saline). Eight subjects from each group (ethanol-naive and chronic-ethanol) received saline injections, and eight subjects from each group received muscimol. Injections were performed over a period of three minutes to minimize tissue damage. Motility was again monitored for 32-second periods, five, ten, 15, 20, and 25 minutes after injection.

Data Analysis For analysis, analog-to-digital conversion of the motility signal and subsequent spectral analysis of the transduced signal were performed with an Apple II microcomputer. A Fast Fourier Transform (Fb'T) was used to obtain power spectra for each one-second segment of motility data, and the spectra were averaged across the 32 seconds of each sampling period. Total power was calculated as the sum of the spectral power measurements for each of the movement frequencies between one and 15 Hz. Following a log transformation of the mean power spectra, a step-wise discriminant function (SDF) analysis was performed to determine the movement frequencies that most contributed to group differences. Univariate analysis of variance with repeated measures was then used to determine the significance of group differences at the various sampling periods with the selected movement frequencies. Duncan's Multiple Range Test was used for individual post hoc group comparisons.

475

9.0 ~" 85]

Q--O NOe'rc~/s~. Q- -0 rroH/s~. 0--0 NO ~O,A~US O- "0 rroH/wUS l

~ ~ ..~

8.o 0

T

T- - 6 - - 7 ~ -

T

•

-- 8:.LL °

7.5,

70. 6.5

•

O1 /

•£

!

I

I

I

I

I

0

,5

10

15

20

2,5

TIME POST-INdECTION (rnin)

FIG. 2. Intranigralmusc]mol-inducedchanges in total power (1-15 Hz) m ethanol-naiveammals(NO-ETOH)and in ammalswithdrawnfrom chronic ethanol exposure (ETOH). Animalsreceivedbilateralintranigralinjectmns of either saline (SAL) or mascimol (MUS). Injections were performed immediatelyfollowingthe motilitymeasurementobtainedat time 0. Total power was obtained as the sum of the spectral power for movement frequenciesbetween 1 and 15 Hz. Data are presented as arbitraryumts of log power___SEM

RESULTS

Mean blood ethanol levels for all of the chronic ethanol animals averaged 368_+28 mg/dl at the time of ethanol withdrawal. Chronic-ethanol animals that subsequently received intranigral saline did not differ significantly in blood ethanol levels at the time of withdrawal from those receiving intranigral muscimol (p>0.05). Ethanol was not detectable in the blood of these animals at the time of behavioral testing 10 hours after withdrawal. At this time, ethanol-withdrawn animals were observed to exhibit whole-body muscle tremors, abnormal posture, and an increased incidence of repetitive sniffing behavior as compared to ethanol-naive controls. SDF analysis of the motility data indicated that prior to muscimol administration, the ethanol-naive and ethanol-withdrawn animals were significantly different from one another (p<0.01). Of the movement frequencies from 1 to 15 Hz, only 9 Hz movements contributed significantly to this difference. In previous studies (8,13), 9 Hz movements have been shown to reflect the occurrence of repetitive, sniffing behavior. Although animals withdrawn from chronic ethanol exposure also exhibited whole-body muscle tremor, the motility monitor was not tuned to be sensitive to this low amplitude, high frequency (>15 Hz) movement (14) and so this behavior was not reflected in the motility data observed. Muscimol administered to ethanol-naive control animals resulted in the occurrence of repetitive, stereotyped movements that were similar to stereotypies observed following the systemic administration of dopamine agonists (31). These animals showed an increased incidence of whole-body movements, vertical head movements, and repetitive sniffing. The observed changes in motility were also consistent with our previous reports of the effects of dopaminergic agonists on motility (13, 16, 17). This included a general increase m power (p<0.0001) at all observed frequencies (1 to 15 Hz), reflecting the observed increase in gross, whole-body movements, with the largest effect of intranigral muscimol seen as an increase in 9 Hz movements. Results of an SDF analysis showed that with the contributions of 9 Hz motility and total power treated as covariates, no other movement frequencies added significantly to the discrimination

between animals receiving intranigral saline and those receiving intranigral muscimol. Since SDF analysis indicated that the effects of both ethanol exposure and muscimol treatment could be best described in terms of 9 Hz motility and total power, subsequent analyses examined just these two variables. All of the groups showed slight, though nonsignificant, increases in total power (Fig. 2) during the first ten minutes following either saline or muscimol injections, perhaps indicating a general behavioral activation as a result of handling during the injection procedure. While the motility of animals receiving intranigral saline then decreased to preinjection levels for the remainder of the observation period, total power (Fig. 2) and 9 Hz motility (Fig. 3) continued to increase in animals that received intranigral muscimol, and these remained high throughout the 25-minute observation period. Analysis of variance indicated that the effects of muscimol on 9 HZ motility and on total power varied significandy with time after muscimol injection (p<0.01). Both ethanol-naive and ethanol-exposed animals receiving muscimol showed significantlymore total power and 9 Hz motility (p<0.01) than did saline-injected animals when tested 15 minutes or later postinjection. Ethanol-exposed animals did not differ from ethanol-naive animals in total power, either before or after intranigral injection. Thus, ethanol-exposed animals showed no significant changes in total power after intranigral saline injection, but did show significant increases (p<0.01) 15 minutes and later following intranigral muscimol, similar to responses observed in ethanol-naive controls. Ethanol exposure and withdrawal did, however, significantly affect 9 Hz motility. Ethanol-exposed animals, which were seen to have significantly more 9 Hz motility than control animals before intranigral injection (p<0.01), also showed significantly more muscimol-induced 9 Hz motility than did control animals receiving saline (p<0.01), but significantly less 9 Hz motility than control animals receiving muscimol (p<0.001). DISCUSSION The results of this study indicate that intranigral administration

476

GONZALEZ AND CZACHURA

7.0-

0--0 O--~

NO E"rOH/SAL

rro./s~_ NO £1"OH/MUS

o--o°--°rro./,.os

"r

I

ol

i

.0-'--- ~

65-

] -0

o-__ 6___ o

w

6.0-

q 55-

5.0

I

I

I

I

I

I

0

5

10

15

20

25

I

TIME POST-INJECTION

(m,.) FIG 3. Nine Hz motility m ethanol-naive animals (NO-ETOH) and in animals withdrawn from chronic ethanol exposure (ETOH). Animals received bilateral lntmmgral injections of either saline (SAL) or muscimol (MUS). Injections were performed immediately following the motility measurement obtained at time 0. Data are presented as arbitrary units of log power--- SEM

of muscimol produced an increase in total power, suggesting a general activation of 1 Hz to 15 Hz movements, and also a specific increase in 9 Hz movements. These observations are consistent with previous reports of stereotypy and general increases in behavior following administration of intranigral muscimol (1,33). Animals chronically exposed to ethanol and withdrawn for ten hours also exhibited increases m total power and in 9 Hz motility after muscimol, as did ethanol-naive controls. The muscimolinduced increase m 9 Hz motility, however, was significantly less m ethanol-withdrawn animals than in ethanol-naive controls. Low doses of intranigral muscimol are reported to induce primarily gross, whole-body movements, with restricted stereotypies observed after higher doses (2). In previous studies of stimulantinduced motility, our laboratory has observed a general increase in the total power of movement frequencies between 1 and 15 Hz associated with an increase in gross, whole-body movements following low doses of stimulant drugs, and a specific increase in 8-10 Hz motility during stereotypy induced by higher doses (13,17). The specific reduction in 9 Hz motility, but not in total power, observed after ethanol withdrawal in the present study, can be interpreted as a shift from restricted stereotypy to general locomotor activation with intranigral muscimol. This reduced behavioral response to intranigral administration of the direct GABA receptor agomst musclmol, suggests that GABA-receptive neurons in the vicinity of the substantia nigra pars reticulata may be functionally less responsive to GABA stimulation during the

period following ethanol withdrawal. This conclusion is m agreement with those of studies examining the effects of withdrawal from chronic ethanol exposure on neurochemical measures of GABA function (34, 35, 40). The GABA receptor complex is thought to consist of at least four binding sites for GABA, picrotoxin, benzodiazepines, and barbiturates (25,41). Chronic ethanol administration does not appear to alter the benzo&azepine or picrotoxin sites, but may affect the coupling of the receptor complex to chloride channels or may alter chloride channels directly (32,38). Other recent studies (5,20) suggest that alterations in GABA function during chronic ethanol exposure might be related to changes in calcium channel activity. This relatively low sensitivity to GABA, together with an increase in calcium channel activity, might result in the neuronal hyperexcitability of the ethanol withdrawal syndrome. An additional possibility is that chronic ethanol exposure could alter activity in other neurotransmitter systems that interact with the nigral GABA system. Activity of the nigrostriatal dopamine system in particular is closely related to nigral GABA function and nigral GABA receptive cells may be important in the mediation of stereotypy induced by dopamme agonists (4,6). Recent research from this laboratory indicates that chronic ethanol administration alters responses to the dopamine agonist apomorphine. These data showed an increase in apomorphine-induced motility (16) and hypothermia (15) following chronic ethanol exposure, suggesting the possibility of increased dopamme-receptor sensitivity. This conclusion is consistent with the finding in the present study that the spontaneous occurrence of 9 Hz motility is increased after chronic ethanol exposure and withdrawal, during the premuscimol period. An alternative explanation of the present data is that ethanol withdrawal may induce the occurrence of behaviors that are incompatible with the occurrence of 9 Hz movements. However, the finding, noted above, that apomorphine-induced 9 Hz movements are actually increased in ethanol-withdrawn animals, indicates that these animals are capable of exhibiting a high incidence of this behavior m spite of the occurrence of other behavioral symptoms of the ethanol withdrawal syndrome. Additional studies will be necessary to examine the possibility of interactions between dopamine-mediated 9 Hz movements and GABA-mediated 9 Hz movements. The present results, together with our earlier studies (15, 16, 18), suggest decreased nigral GABA responsiveness at the same times relative to ethanol withdrawal when animals show increased responsiveness to dopaminergic stimulation and when they exhibit a variety of symptoms of the ethanol withdrawal syndrome, including sensitivity to audiogenic seizures. Studies are currently in progress in this laboratory to determine if the changes in functional sensitivity that we have observed to dopamine and GABA stimulation during ethanol withdrawal are the result of direct changes in neuronal receptor responsiveness.

REFERENCES 1. Arnt, J., Scheel-Kruger, J Behavioral differences reduced by muscimol selectively injected into pars compacta and pars reUculata of the substantia mgra. Arch Pharmacol 310:43-51, 1979 2. Bachus, S. E., Gale, K Differential effects of chronic treatment with chlorpromazine versus cocaine on behavioral responsiveness to mgral GABA receptor stimulation. Psychopharmacology (Berlin) 95'56-62, 1988 3 Dar, M. S., Wooles, W R GABA mediation of the central effects of acute and chronic ethanol m mice. Pharmacol. Blochem Behav 22(1) 77-84; 1985 4 DeMontas, G., Ohanas, M. C , Serra, G , Taghamonte, A., ScheelKruger, J. Evidence that a mgral GABAergic-cholinerg]c balance controls posture. Eur J Pharmacol 53 181-190, 1979.

5 Dohn, S ; Little, H., Hudsplth, M , Pagoms, C , Llttleton, J Increased dihydropyndine-sensitave calcium channels m rat brain may underlie ethanol physical dependence Neurophannacology 26(23):275-279; 1987 6. Dray, A, The smatum and substantla mgra. A commentary on their relationships. Neurosclence 4:1407-1439, 1979. 7. Elhnwood, E H., Jr., Kllbey, M. M Alteration in motor frequencies with dopamane agomsts and antagonists. Psychopharrnacol Bull. 15:49-50; 1979. 8 Ellinwood, E H., Jr., Molter, D. W , Stauderman, K A. An assessment of the spectral analysis of amphetanune-mduced behavior Pharmacol. Blochem. Behav 15.627--631, 1981. 9 Frye, G D., Breese, G R GABAerglc modulauon of ethanol-

CHRONIC ETHANOL AND INTRANIGRAL MUSCIMOL

10.

11. 12.

13. 14. 15.

16 17.

18. 19. 20.

21.

22. 23.

24. 25.

26. 27.

reduced motor impairment. J. Pharmacol Exp. Tber. 223:750-756, 1982. Frye, G. D.; McCown, T. J.; Breese, G. R. Differential sensaway of ethanol withdrawal signs in the rat to "~/-aminobutyric acid (GABA)mimetics. Blockade of androgenic seizures but not forehmb tremors. J. Pharmacol Exp Ther. 226(3):720-725; 1983. Goldstem, D. B. Sodium bromide and sodium valproate: Effect|ve suppressants of ethanol withdrawal reactaons in mice. J. Pharmacol. Exp. Ther. 208(2):223-227; 1979. Goldstem, D. B. Alcohol withdrawal reactaons m mice- Effects of drugs that modify neurotransmission J. Pharmacol. Exp. Ther 186:1-9; 1973 Gonzalez, L. P. Alterations in amphetamine stereotypy following acute lesions of substantm nlgra. Life SCL 40:899-908, 1987. Gonzalez, L. P. Quantitative analysis of physostlgmine-lnduced changes m behavior Pharmacol. Bmchem Behav. 21'551-554, 1984 Gonzalez, L. P , Colbern, D.; Czachura, J. F. Apomorphme-mduced hypothermia increases during ethanol withdrawal. Alcohol 5 107109, 1988 Gonzalez, L. P., Czachura, J. F. Ethanol withdrawal alters apomorphme-induced motlhty. Pharmacol Biochem. Behav. 31:163-168, 1988. Gonzalez, L. P , Ellinwood, E. H., Jr. Cholinerglc modulatmn of stimulant-induced behavior Pharmacol Blochem. Behav. 20:397403, 1984. Gonzalez, L P.; He~nger, M. K. Intranigral musclmol suppresses ethanol withdrawal seizures. Brain Res. 298 163-166, 1984. Haklonen, H.-M.; Kulonen, E. Ethanol mtoxtcataon and -/-amlnobutync acid. J. Neurochem. 27'631-633, 1976. Hudspith, M. J., Brennan, C H., Charles, S., Littleton, J. M. Dthydropyndme-sensitive CA 2+ channels and inosltol phosphohpld metabohsm in ethanol physical dependence. Ann. NY Acad SCL 492:156-170; 1987. Ito, M. Roles of GABA neurons in integrated functions of the vertebrate CNS. In: Roberts, E , Chase, T. N , Tower, D. B., eds. GABA in nervous system function New York: Raven Press, 1976 427. Leltch, G J., Backer, D. J , Siegman, F. S.; Guthne, G D Possible role of GABA m the development of tolerance to alcohol. Experientla 33:496--498; 1977. Lfljequlst, S.; Engel, J Effects of GABAerglc agomsts and antagomsts on various ethanol-induced behaworal changes Psychopharmacology (Berlin) 78:71-75, 1982. Majchrowlcz, E. Induction of physical dependence on alcohol and the assocmted behavioral changes m rats. Psychopharmacologla 43.245254; 1975. Maksay, G., Ticku, M. K. Dissociation of (35S)t-butylblcylcophosphorothlonate binding differentiates convulsant and depressant drugs that modulate GABAergic transmission. J. Neurochem 44'480--486, 1985. Martz, A., Deitrich, R. A.; Hams, R. A. Behavioral evidence for the involvement of "y-aminobutync acid in the actions of ethanol Eur. J. Pharmacol 89'53-62; 1983 Nestoros, J. N. Ethanol specifically potentiates GABA-medlated neurotransmassinn m fehne cerebral cortex Science 209.708--710, 1980.

477

28 Nestoros, J. N. Ethanol selectively potentiates GABA-mediated inhibition of single fehne cortical neurons. Life Scl. 26.519-523; 1980. 29. Nordberg, A , Wahlstrom, G., Eriksson, B. Relations between musclmol, qumuchchnyl benzilate and nicotine binding in brain after very long treatment with ethanol in rats. Eur J. Pharmacol. 115 301-304; 1985. 30. Paxinos, G., Watson, C. The rat brain m stereotaxic coordinates. New York: Acadenuc Press; 1982. 31. Randrup, A., Munkvad, I. Pharmacology and physiology of stereotyped behavior. J Psychiatry Res. 11.1-10; 1974. 32. Rastogl, S. K.; Thyagarajan, R., Clothier, J.; Ticku, M. K. Effect of chromc treatment of ethanol on benzodiazepme and picrotoxin sites on the GABA receptor complex in regions of the brmn of the rat Neuropharmacology 25(10):1179-1184, 1986. 33 Scheel-Kruger, J , Arnt, J.; Magelund, G. Behavioral stirnulaUon induced by musclmol and other GABA agomsts rejected into the substantaa mgra. Neuroscl. Lett. 4 351-356; 1977. 34. Simler, S., Clement, J.; Cies~elslo, L.; Mandel, P. Brain ~/-aminobutync acid turnover rates after spontaneous chronic ethanol intake and withdrawal m discrete brain areas of C57 rmce. J. Neurochem 47:1942-1947, 1986. 35. Sytlnsky, I. A.; Guzikov, B M.; Gomanko, M. V ; Eremin, V P , Konovalova, N N. The gamma-aminobutync acid (GABA) system m brain dunng acute and chronic ethanol intoxication. J. Neurochem 25 43-48; 1975 36. Taberner, P. V.; Unwin, J W. Behavioural effects of musclmol, amphetamme, and chlorpromazine on ethanol tolerant mace. Br. J. Pharmacol. 74.2761-2762; 1981. 37. Tanka, J S., Winger, G The effects of ethanol, phenobarbital, and baclofen on ethanol withdrawal In the rhesus monkey Psychopharmacology (Berlin) 70'201-208, 1980. 38 Thyagarajan, R., Tlcku, M. K. The effect of m v,tro and in vivo ethanol administration on (3~S)t-butylbicyclophosphorothlonate binding in C57 mace. Brain Res. Bull. 15:343-345, 1985. 39 Tlcku, M. K The effects of acute and chrome ethanol admmlsta'atlon and its wahdrawal on gamma-anunobutyric acid receptor binding Br. J Pharmacol. 70"403-410, 1980. 40 Tlcku, M. K , Birch, T. Alterations m ~-amanobutyric acid receptor sens~ttvity following acute and chromc ethanol treatments. J Neurochem 34(2):417-423, 1980. 41. Tlcku, M. K , Rastogl, S K. Barbiturate-sensitive sites in the benzothazeplne-GABA receptor-ionophore complex. In" Roth, S. H.; Mdler, W. K., eds. Molecular and cellular mechanisms of anesthetics New York: Plenum Press, 1980:179-188. 42. Unwln, J. W , Taberner, P. V. Sex and stram differences in GABA receptor binding after chromc ethanol dlrnking m mice. Neuropharmacology 19.1257-1259, 1980 43 Volicer, L.; Blagioni, T. M. Effect of ethanol admimstrataon and withdrawal on GABA receptor binding m rat cerebral cortex. Subst. Alcohol Actaons/Misuse 3 31-39; 1982. 44 Wlxon, H. N., Hunt, W. A. Effect of acute and chromc ethanol treatment on gamma-anunobutync acid levels and on armnooxyacetlc acid-reduced GABA accumulation Subst. Alcohol Actaons/Misuse 1:481-491, 1980.