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Antagonism of the effects of the atypical benzodiazepine, Ro 5-4864 on intracranial self-stimulation in the rat
Phurmu
f?wthenzr\w\
& Rrha~ror,
Vol 24. pp 193-197. 1986 T Ankho InternatIonal
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Antagonism of the Effects of the Atypical Benzodiazepine, Ro 5-4864 on Intracranial Self-Stimulation in the Rat SHARON
L J HERBERG*
PELLOW,’
AND SANDRA
E. FILE
MRC Nrrrrophurmut ology Rcsc)urc h Group Unr\lcrsrty of London. 29-39 Bruns\tslc h Square, London WCIN IAX, U K *E.xpcr~m~~ntulPryc hologv Luhoratoty, In ttltntc of Neurology, Queen Squaw, London WC’IN 3BG. U K Recewed
25 March
1985
PELLOW. S . L J HERBERG AND S E FILE Antagonrw~ of rhr cqjcc t\ oj rhc LII\~ICtrl hen~odicrzeprrrf, Ro 5-4864 on rrr[ruc rcr~rrul \clf-,/rrrrr,lofrotl,n /hc~rut PHARMACOL BIOCHEM BEHAV 24(Z) 193-197, 1986 -The effects of Ro 5-4864 (chlordlazepam) were exammed on respondmg for self-stunulation of the mid-lateral hypothalamus Rewardmg stimuli were delivered accordmg to a IO-set varlable Interval schedule of remforcement Ro S-4864 (l&30 mg/kg, subconsulslve dotes m these rats) decreased responding Thts effect was antagomzed by chlordtazepoxtde (5-10 mg/kg) and phenobarbltone (35 mg/kg) but not by the benzodlazepme receptor antagomst Ro 15-1788 (l&20 mg/kg), the hgand for penpheral benzodlazepme receptors PK I1 195 (60 mg/kg) or phenytom (60 mgikg) The pattern of mteractlons of Ro 5-4864 with these compounds differs from the pattern obtatned with other procedures, and suggests that Ro 5-4864 has effects on systems unrelated to anxiety, convulsive activity or sedatton Intracramal
self-stlmulatlon
Ro S-4864
Benzodlazepme
Ro 5-4864 15 a 1,4-benzodlazepme that, although havmg neghglble affmlty for ‘classical’ CNS benzodlazepme receptors, bmds with high affmlty to ‘peripheral-type’ benzodlazepme bmdmg sites that are present m the periphery [4] and m the CNS 1271 Although Ro 5-4864 has no benzodlazepme-hke actlvlty m ammal tests [31] and thus has long been consldered mactlve [24], recent mvestlgatlons mto Its behavloural effects have shown that It has convulsant [9, 22, 25, 301, anxlogemc [S. I I, 12, 191 and sedative [lo] propertles Further behavloural. electrophyslologlcal and blochemlcal mvestlgatlons have questioned the conclusion, made on the basis of bmdmg studies, that Ro 5-4864 does not Interact with the GABAlbenzodlazepmellonophore complex m the CNS, and suggest a possible action at the plcrotoxmm site that IS linked with the chlonde lonophore on this complex (see [ 19,201 for revtews) Respondmg for lateral hypothalamic self-stlmulatlon on a vanable Interval schedule of remforcement has been shown to be a sensltlve procedure for ldentlfymg and dlstmgmshmg the behavloural effects of benzodlazepmes, GABA agonists and antagonists, non-GABAerglc antlconvulsants [ 131 and benzodlazepme ‘antagomsts such as Ro 15-1788, CGS 8216 and FG 7142 [23,24] Our aim was to charactenze further the behavloural effects of Ro 5-4864 usmg this procedure, and to examme Its profile m combmatlon with several other compounds with which It has previously been shown to interact These
were
PK
I 119.5, an tsoqutnolme
‘Correspondence at the followmg address boulevard de I’HBpltal. 75634 Pans CCdex
carboxamtde Department 13, France
antagonism
tlve that, hke Ro 5-4864, selectively bmds to penpheral-type benzodlazepme bmdmg sites [ 15,161 and 1s thought to be an antagonist there [ 171, chlordlazepoxlde, a conventional I ,4 benzodlazepme with moderate potency for classlcal CNS benzodlazepme receptors but with neghglble affimty for penpheral-type sites [27], Ro 15-1788, an lmldazodlazepme that antagonizes several of the effects of benzodlazepmes and p-carbolmes [ 14,211 and that IS beheved to interact only with classlcal CNS benzodlazepme receptors [26], phenobarbltone, a barbiturate that displaces [‘HI-adlhydroplcrotoxmm and [ %S]-t-butyl blcyclophosphorothlonate from the plcrotoxm site, and phenytom, which Interacts with benzodlazepmes at low affinity bmdmg sites m the brain [2, 3,7, 18, 281 Doses of chlordlazepoxlde, Ro 15-1788, phenobarbltone and phenytom were chosen on the basis of previous studies ([13,23], Pellow and File, unpubhshed), dose-response data were determmed for Ro 5-4864 and PK I I I95 METHOD AnlnlUlJ
Male hooded PVG rats (Bantm and Kmgman Ltd ) weighing 230-250 g at time of surgery were mdlvldually housed with free access to food and water Under halothane anesthesia, twisted bipolar stainless steel electrodes (0 25 mm diameter, Plastic Products Inc ,
dertvade Pharmacologle.
193
Faculte
de Medecme,
Pltl&SalpCtniire,
Umverslte
Pan\
VI. 91
194
PELLOW, HERBERG AND FILE TABLE 1 MEAN ( + S E M ) S E L F - S T I M U L A T I O N R A T E (% B A S E L I N E ) IN E A C H 15-MIN PERIOD OF A 60-MIN TEST SESSION, IN RATS TESTED I M M E D I A T E L Y A F T E R IP INJECTION WITH Ro 5-4864 (10 AND 30 mg/kg) OR PK 11195 ( 1 0 ~ 0 mg/kg)
Drug
1st 15-mm
2nd 15-mm
3rd 15-mm
4th 15-mm
control Rol0 Ro30
982 _+ 1 03 952_+251 728_+ 6 16"
998 _+ 1 67 9 2 6 _+ 377 68 1 -+ 656"
975 _+ 3 61 952_+ 455 589_+ 104"
101 3 _+ 4 17 995_+ 503 634_+ 108"
PK 10 PK 30 PK60
952_+ 443 928 + 3 75 874-+ 723
997_+ 459 91 8 -+ 13 4 894_+ 750
101 5_+ 697 984 + 3 43 869_+ 800
962_+ 10 10 1006-+ 808 955 _+ 12 20
*p<0 05
R o a n o k e , VA) were stereotaxlcally implanted in the midlateral hypothalamus 1 0 m m posterior to Bregma, 1 3 mm to the left of the m,dllne and 8 0 m m below the skull surface E l e c t r o d e placements w e r e verified on enlarged photographic projects of unstained 50-/am frozen sections at the end of the investigation
150 i
125
"6 P
100
CONTROL
Ro 5-4864 (301
"/5
Drug~ Ro 5-4864 ( H o f f m a n - L a Roche), P K 11195 (Pharmuka), p h e n y t o m (Sigma), sodium p h e n o b a r b l t o n e ( B D H Chemicals) and Ro 15-1788 ( H o f f m a n - L a Roche) were suspended m distilled water with a drop of T w e e n 20 and dispersed by ultrasound, c h l o r d l a z e p o x l d e h y d r o c h l o n d e (Roche Products) was dissolved in distilled water Control injections consisted of dlstdled water, with or without a drop of T w e e n 20, as appropriate Drugs were injected ,ntrapentoneally ,n c o n c e n t r a t i o n s to give an rejection v o l u m e of 2 ml/kg
SelJ-Sttmulatton Rats were allowed to r e c o v e r from surgery for a week, and then trained to operate a pedal for a 0 5-sec 50-Ha s m e w a v e r e m f o r c m g impulse available on a variable interval schedule o f 10-sec m e a n duration (see [13, 23, 24] The stimulating current for each rat was fixed at the lowest intensity that elicited uninterrupted responding in preliminary trials in which intensities were d e c r e m e n t e d m decdog steps, b e t w e e n 50 and 10/xA r m s R e s p o n s e rates in different rats ranged b e t w e e n 15 and 25 l e v e r - p r e s s e s / m m During test sessions, animals w e r e allowed to selfstimulate for 45 mm, of which the last 30 mln provided a pre-mjectlon basehne The rat was then rejected, and allowed to respond for a further 75 mln If the rat stopped responding it was encouraged to restart by taps on the Fever, if this failed, by admlmstratlon of priming shocks, and, finally, by the e x p e r i m e n t e r placing the rat bodily on the lever R e s p o n s e rates w e r e r e c o r d e d automatically at 5-mm intervals as a dlg,tal printout Drug effects w e r e determined from the rate recorded during each 15-mln period following mjecUon, e x p r e s s e d as a percentage of pre-mjection basehne, and c o m p a r e d with the corresponding rate after control injections P/o( edut e
Expertment I Dose response data were obtained for Ro 5-4864 and PK 11195 on self-stimulation responding Drug
5O
_e 25
~,
o
Control PK CDP Pheny- Control PK CDP Pheny11195 (.5) O0) tom 11195 (5) (lO} tom (60) (60) (60} (60)
FIG 1 Mean (+S E M ) self-stlmulatzon rate (variable interval l0 sec) m the second 15-ram period after injection for rats given phenytom (60 rng/kg) or chlord,azepoxide (5 or 10 mg/kg) alone or m cornbmatlon with Ro 5-4864 (30 mg/kg)
groups were vehicle, Ro 5-4864 10 and 30 mg/kg, PK I 1195 10, 30 and 60 mg/kg ( n = 7 per group) L~pertment 2 The effects of an active dose of Ro 5-4864 (30 mg/kg) were investigated in combination wlth p h e n y t o m (60 mg/kg), p h e n o b a r b , t o n e (35 mg/kg), Ro 15-1788 (10 and 20 mg/kg), chlordlazepoMde (5 and 10 mg/kg), PK 11195 (60 mg/kg) ( n = 6 per group) Each rat was tested at 48-hr intervals, and m each exper,ment the drug treatments were administered m a predetermined random o r d e r that was different for each rat Statl~tt~ s
Data from E x p e r i m e n t 1 were analysed by Friedman 2-way analysis of variance, and c o m p a r i s o n s between individual groups were made with W d c o x o n matched-paws signed-ranks tests Data from E x p e r i m e n t 2 were analysed by W d c o x o n matched-pairs signed-ranks tests RESULTS
E~permwnt I Ro 5-4864 (10-30 mg/kg) caused a slgmficant dose-related reduction m responding for self-stimulation m the first, X-'=6 0, p < 0 05, second, X-'= 12 2 9 , p < 0 005, third, X-'=8 86, p < 0 05, and fourth, X-'= 10 57, p < 0 005, 15-mm time periods after injection (see Table 1) W d c o x o n tests showed that at 30 mg/kg slgmficant reductions m self-sumulatlon respond-
A N T A G O N I S M O F E F F E C T S OF Ro 5-4864
195 TABLE 2
S U M M A R Y T A B L E OF T H E ABILITY O F S E V E R A L C O M P O U N D S TO A N T A G O N I S E T H E E F F E C T S O F Ro 5-4864 IN A N I M A L TESTS O F ANXIETY, C O N V U L S I V E ACTIVITY, E X P L O R A T I O N A N D S E D A T I O N , A N D IN T H E P R E S E N T S E L F - S T I M U L A T I O N P R O C E D U R E
Drug Benzodmzepmes Barbiturates Ro 15-1788 PK 11195 Phenytom
Anxiety ~ [111 ~* - [Ill - [8,12] ~ [8]
Convulsions
Exploration
~ [7,29] ~ [291 ~ [71 $ [6,8] ~ [8]
Motor
~ ~ - [10l $ [10] - [10]
') ') - [101 - [10] [10]
ICSS
-
*Pellow and Fde, unpubhshed data compound antagomses Ro 5-4864, - compound does not affect Ro 5-4864, '~ not yet reported References to studies are in brackets (see text)
mg were observed at all time-points PK 11195 had no s)gmf)cant effect at any t~me period, X-'=2 49, 2 48, 0 09, 0 43, respectwely), see Table 1
150
CONTROL
Ro 5-4864 (30)
lz5
I~ ~ p e r t m e n t 2
"5 100
Phenytom (60 mg/kg) s)gmficantly decreased selfst~mulat)on m the second 15-mm period after mjecUon (p<0 05) but d)d not antagonize the reduction produced by Ro 5-4864 (30 mg/kg) (see F~g 1) CDP increased selfst)mulat)on at 5 mg/kg (p<0 05) and decreased )t at 10 mg/kg (p<0 05), and at both doses s~gmficantly reversed the reduct)on produced by Ro 5-4864 (30 mg/kg, p < 0 05, see F~g 1) Interestingly, however, Ro 5-4864 d~d not antagomze the mcrease m self-stimulation produced by CDP (5 mg/kg) PK 11195 (60 mg/kg) had no s)gmficant effect on self-st)mulaUon alone, and d~d not antagomze the effects of Ro 5-4864 (see F)g 2) Ro 15-1788 (10-20 mg/kg) d)d not antagomze the effects of Ro 5-4864, although at 20 mg/kg th)s compound when g)ven alone s)gmficantly increased self-sUmulaUon (see F)g 2) Phenobarb)tone (35 mg/kg) s)gmficantly increased self-st~mulat)on (p<0 05), and also antagomzed the reduced self-sUmulaUon produced by Ro 5-4864 (30 mg/kg, p < 0 05, see F~g 2) DISCUSSION
Ro 5-4864 at 30 mg/kg produced a decrease m responding for lateral hypothalam)c self-st~mulauon, s,mflar to that reported for the benzodmzepme receptor 'reverse agomsts' CGS 8216 and FG 7142 [23.24] Since Ro 5-4864 is a compound w)th known convulsant acuons, ,t ~s necessary to cons)der the extent to wh)ch these act)ons could account for )ts effects on self-stimulation The dose of Ro 5-4864 that has been found to effect,rely cause convulsions has vaned conslderably, both between (c f [1,6, 25, 29]) and within (cf [6, 7, 22]) laboratories In our expenence, d)fferences have been observed between species (cf. [6,7]) and stratus (cf [6,9]), and also between different batches of the supphed compound In each new batch of experiments, then. we have found ,t necessary to estabhsh the effective convulsant dose of Ro 5-4864 before carrying out the intended behavloural work During the course of the present study we observed, m th,s strain of rats, that convulsions did not occur at doses below 90 mg/kg At 90 mg/kg 100% rats convulsed, at 60 mg/kg we only observed myoclomc spasms m one rat who had been treated and tested for self-st~mulaUon responding In no cases were convulsant-hke activities observed at 30
75 -5
Control
15-1788 Pheno (10) (20) barb (35)
Control
15-1788 Pheno(10) (20) barb (35)
FIG 2 Mean (=S E M ) self-sumulatJon rate (variable interval, 10 sec) m the second 15-mm period after rejection for rats given PK 11195 (60 mg/kg) and Ro 15-1788 (10 or 20 mg/kg) alone or m comblnaUon with Ro 5-4864 (30 mg/kg)
mg/kg It is therefore unlikely that the convulsant properties of Ro 5-4864 were contributing to ~ts effects on selfst~mulauon The pattern of mteracUons of Ro 5-4864 w~th other drugs (see below and Table 2) supports this view PK 11195 alone (10-60 mg/kg) had no stgmficant effect on self-st~mulaUon, htgher doses were not investigated since at the 60 mg/kg dose some ammals showed unusual body postures that could have interfered w~th their ablhty to lever-press The reducUon m self-stimulation produced by Ro 5-4864 (30 mg/kg) was not antagomzed by PK 11195 (60 mg/kg), this is consmtent with the inability of PK 11195 to antagomze the effects of Ro 5-4864 m the socml Interaction and pumshed drmkmg procedures [12]. although PK 11195 does antagomze the proconvulsant and convulsant effects [1,9] and the reductions m exploratory head-dipping produced by Ro 5-4864 [10] As found previously [13, 23, 24], chlordlazepoxlde (5 but not 10 mg/kg) Increased self-stimulation Chlordmzepoxlde at both doses sJgmficantly reversed the reducuon m response rates produced by Ro 5-4864, however, interestingly, Ro 5-4864 was unable to antagomze the increase m selfstimulation produced by chlordmzepoxlde (5 mg/kg) Chlordmzepox~de is able to antagomze the anxlogemc effects of Ro 5-4864 [11], and other benzodmzepmes have been found to antagomze the convulsant effects of th~s compound [7,29] Th~s )s s~mdar to the effects obtained by Herberg and Wllhams [13] w)th plcrotoxm and chlordmzepoxlde, however, unhke plcrotoxm. Ro 5-4864 did not produce a further
196 e n h a n c e m e n t o f r e s p o n d i n g w h e n c o m b i n e d with chlord l a z e p o x l d e (10 mg/kg) R o 15-1788 (20 mg/kg) e n h a n c e d self-stimulation, as previously f o u n d b y P e | l o w et a! [23] This c o m p o u n d did n o t a n t a g o n i z e t h e r e d u c t i o n in s e l f - s t i m u l a t i o n p r o d u c e d b y Ro 5-4864, again c o n s i s t e n t with findings in t h e social i n t e r a c tion t e s t o f a n x i e t y a n d the h o l e b o a r d [10,11] H o w e v e r , R o 15-1788 (20 mg/kg) IS able to a n t a g o n i z e the c o n v u l s i o n s p r o d u c e d b y R o 5-4864 in at least s o m e c i r c u m s t a n c e s [7] P h e n o b a r b l t o n e (35 mg/kg), like the b a r b i t u r a t e p h e n o b a r b l t o n e [13], i n c r e a s e d s e l f - s t i m u l a t i o n a n d a n t a g o n i z e d t h e effects o f Ro 5-4864 Again, similar r e s u l t s h a v e b e e n obt a i n e d m t h e social i n t e r a c t i o n test (Pellow a n d File, u n p u b h s h e d ) , w h e r e a d o s e o f p h e n o b a r b l t o n e t h a t also i n c r e a s e d social m t e r a c t i o n w a s able to r e v e r s e the d e c r e a s e p r o d u c e d b y R o 5-4864 H o w e v e r , it is n o t p o s s i b l e to be sure w h e t h e r t h e a n t a g o n i s m b e t w e e n t h e s e t w o c o m p o u n d s in b o t h of t h e s e t e s t s is specific o r w h e t h e r it simply reflects a cancell a t i o n o f t w o o p p o s i t e intrinsic a c t i o n s B a r b i t u r a t e s also p r e v e n t the c o n v u l s i o n s usually o b s e r v e d with Ro 5-4864 [29] C o n s i s t e n t w i t h t h e r e p o r t of H e r b e r g a n d Williams [ 13], p h e n y t o l n d e c r e a s e d s e l f - s t i m u l a t i o n T h e Inability of p h e n y toin (60 mg/kg) to a n t a g o n i z e the d e c r e a s e in s e l f - s t i m u l a t i o n p r o d u c e d b y R o 5-4864 c o n t r a s t s with its ability to antagonize the a n x l o g e n l c a n d c o n v u l s a n t a c t i o n s o f this c o m p o u n d [8,9] b u t IS similar to its inability to a n t a g o n i z e t h e r e d u c h o n s
PELLOW, HERBERG AND FILE in e x p l o r a t o r y head-d~pplng a n d l o c o m o t o r activity in the h o l e b o a r d p r o d u c e d b y R o 5-4864 [10] It ~s unlikely t h a t the d e p r e s s a n t effect that p h e n y t o i n p r o d u c e s w h e n given a l o n e w o u l d interfere with its ability to a n t a g o n i z e the r e d u c t i o n p r o d u c e d by Ro 5-4864, since c h i o r d m z e p o x l d e at a d o s e t h a t d e c r e a s e d r e s p o n s e r a t e s (10 mg/kg) was stdl able to antagonize the r e d u c t i o n p r o d u c e d b y Ro 5-4864 In c o n c l u s i o n , the p a t t e r n of i n t e r a c t i o n of Ro 5-4864 with o t h e r c o m p o u n d s m the v a r i a b l e - i n t e r v a l self-stimulation p r o c e d u r e s h o w s a b r o a d similarity to the p a t t e r n s s e e n in v a r i o u s o t h e r p r o c e d u r e s , b u t t h e r e are also s o m e n o t a b l e d i f f e r e n c e s (see T a b l e 2) It ~s t h u s u n c e r t a i n w h e t h e r the effects o f Ro 5-4864 in this p r o c e d u r e are to be a t t r i b u t e d to its effects o n a n x i e t y , c o n v u l s a n t activity or s e d a t i o n , or to ~ts effects on o t h e r , u n r e l a t e d s y s t e m s T h e s a m e q u e s t i o n applies to s e v e r a l related a g e n t s t h a t h a v e also b e e n f o u n d to d e p r e s s s e l f - s t i m u l a t i o n , s u c h as the b e n z o d m z e p m e recept o r ' i n v e r s e a g o n l s t s ' F G 7142 a n d C G S 8216, a n d the recept o r a n t a g o n i s t Ro 15-1788 [23.24] E x p e r i m e n t s are c u r r e n t l y u n d e r w a y to i n v e s t i g a t e th~s q u e s t i o n
ACKNOWLEDGEMENTS SEF ~s a Wellcome Trust Semor Lecturer We are grateful to the Medical Research Councd for eqmpment
REFERENCES 1 Benawdes, J , F Gullloux, D E Allam, A Uzan, J Mlzoule, 13 Herberg, L J and S F Wdhams Ant~confl~ct and depressant effects by GABA agonists and antagomsts, benzodlazepmes and C Renault, M C Dubroeucq, C Gueremy and G Le Fur non-GABAerglc antlconvulsants on self-stimulation and locoOpposite effects of an agomst, Ro 5-4864, and an antagomst, PK motor activity Pharma~ol Btoc hem Behm 19: 625-633, 1983 11195, of the peripheral type benzodmzepme binding sites on 14 Hunkeler, W , H Mohler, L Plen. P Polc, E P Bonettl R audlogemc seizures m DBA/2J mice Life S~; 34" 2613-2620, Cumin, R Schaffner and W Haefely Selective antagonists of 1984 benzodlazeplnes Nature 290: 514-516, 1981 2 Bowhng, A C and R J DeLorenzo Micromolar affimty ben15 LeFur, G , F Gudloux, P Rufat, J Benavldes, A Uzan, C zodlazeplne receptors identification and characterlsatlon m Renault, M C Dubroeucq and C Gueremy Peripheral benzocentral nervous system St wnce 216" 1247-1249, 1982 dlazeplne binding s~tes effect of PK 11195, I-(2-chlorophenyl)3 Bowhng, A C and R J DeLorenzo Multiple phenytom bradN-methyl-N-(I-methylpropyl)-3-1soqumohne carboxamlde II In mg sites ldentlficatmn and charactensaUon m brain membrane vlvo studies Lth" 3~t 32. 1849-1856, 1983 So~ Neuros~t Abstr 8. 505, 1982 16 LeFur, G , M L Pemer, N Vaucher, F Imbault, A Flamler, 4 Braestrup, C and R F Sqmres Specific benzodmzepme recepJ Benavldes. A Uzan, C Renault, M C Dubroeucq and C tors m rat brain characterized by high-affinity [3H]-dmzepam Gueremy Peripheral benzodlazeptne binding sites effect of PK binding Pro~ Natl Acad S~t USA 74: 3805-3809, 1977 11195, l-(2-chlorophenyl)-N-methyl-N-( l-methylpropyl)-35 Burnham, W M , L Spero, M M Okazakland B K Madras isoqumollne carboxamlde I In wtro studies Lt/e St; 32 1839Saturable binding of [3H]-phenytom to rat brain membrane 1847. 1983 fractmns Can J Physlol Pharma~ ol 59. 402-408, 1981 17 LeFur, G . N Vaucher. M L Perner, A Flamler, J Ben6 File, S E Pro- and ant~-convulsant properties of PK 11195, a awdes, C Renault, M C Dubroeucq C Gueremy and A hgand for benzodmzepme binding sites development of Uzan Differentiation between two hgands for peripheral bentolerance Br J Pharmacol 83: 471-476, 1984 zodlazeplne binding sites, [ ~H]-Ro 5-4864 and [ ~H]-PK I 1195, by 7 Fde, S E , A R Green, D J N u t t a n d N D Vincent On the thermodynamic studies Life S~t 33: 44%457, 1983 convulsant actmn of Ro 5-4864 and the existence of a m~cromo18 Okazakl, M M , B K Madras, K E Llwngston, L Speroand lar benzodmzepme binding s~te m rat brain Psv~ hopharma~ olW M Burnham Enhancement of [~H}-phenytom binding by ogv (Berhn) 82: 19%202, 1984 dmzepam and bJcuculhne Life S~I 33. 409-414, 1983 8 File, S E and R G L~ster The anxlogemc action of Ro 5-4864 19 Pellow, S and S E Fde Characteristics of an atypical benis reversed by phenytom Neuros~t Lett 35" 93-96, 1983 zodmzepme, Ro 5-4864 Neuro~l Bu~behm Re~ 8 405-413, 9 File, S E and P S Mabbutt Behav~oural effects of Ro 5-4864, 1984 a hgand for the m~cromolar benzodmzepme receptor Br J 20 Pellow. S and S E File Behawoural effects of Ro 5-4864 a Pharma~ol 78: 76P, 1983 peripheral-type benzodmzepme "~Life St t 35" 22%240, 1984 10 Fde, S E and S Pellow Ro 5-4864, a hgand for benzodmzepme 21 Pellow, S and S E Fde Multiple sites of action for anxtogemc m~cromolar and peripheral binding sRes antagonism and drugs behawoural, electrophysmloglcal and biochemical correenhancement of behavmural effects Psy~hopharrna~ ology lations Psw hopharma~ olog~ (Berhn) 83:304-315, 1984 (Berhn) 80: 166-170, 1983 22 Pellow, S and S E Fde Proconvulsant and antlconvulsant 11 File, S E and S Pellow The anxmgemc actmn of Ro 5-4864 drug effects m combination w~th the convulsant benzodmzepme effect of chlordlazepox~de, Ro 15-1788 and CGS 8216 Naunvn Ro 5-4864 I PhaHn Pharma(ol 37" 560-563, 1985 S~hmwdebergs Arc h Pharma~ ol 328: 225-228, 1985 23 Pellow, S , S E File and L J Herberg lntracranml self12 File, S E and S Pellow The effects of PK 11195, a hgand for stimulation d~stlngmshes between two benzodlazepme benzodmzeplne binding sites, in ammal tests of anxiety and antagomsts Neuros~t Lett 47" 173-177. 1984 stress Pharmat ol Bto~hem Behav, in press, 1985
A N T A G O N I S M O F E F F E C T S O F Ro 5-4864
24 Pellow, S , L J Herberg and S E Fde The effects of the /3-carbohne, FG 7142, on mtracranml self-stimulation in the rat Pharmac ol Bto~ hem Behav 21: 667-669. 1984 25 Plen, L , P Polc, E P BonettL W Burkard, R Cumin, R Scherschhcht and W Haefely Some pharmacologxcal effects of Ro 5-4864, a specific hgand for the peripheral type of benzodlazepme binding sites N a u n v n S( hmtedeberg~ Arc h Pharma
197
28 Shah, D S , P Chambon and A Guldottl Binding of[~H]-5,5 dlphenylhydantoln to rat brain membranes Neuropharma~ ologv 20. 1115-1119 1981 29 Welssman, B A , J Cott, D Hommer, S Paul and P Skolmck Electrophyslologlcal and pharmacological actions of the convulsant benzodlazepme Ro 5-4864 Lttr I Pharma~ ol 97. 257263 1984 30 Welssman, B A , J Cott, S M Paul and P Skolmck Ro 5-4864 a potent benzodlazepme convulsant Lur I Pharma~ ol 90. 149-150, 1983 31 Zbmden, G and L O Randall Pharmacology of benzodlazepmes laboratory and chmcal correlations Ad~ Pharmayo/5 213-291, 1967
f?wthenzr\w\
& Rrha~ror,
Vol 24. pp 193-197. 1986 T Ankho InternatIonal
Inc Pnnted III the U S A
0091-3057186 $3 00 + 00
Antagonism of the Effects of the Atypical Benzodiazepine, Ro 5-4864 on Intracranial Self-Stimulation in the Rat SHARON
L J HERBERG*
PELLOW,’
AND SANDRA
E. FILE
MRC Nrrrrophurmut ology Rcsc)urc h Group Unr\lcrsrty of London. 29-39 Bruns\tslc h Square, London WCIN IAX, U K *E.xpcr~m~~ntulPryc hologv Luhoratoty, In ttltntc of Neurology, Queen Squaw, London WC’IN 3BG. U K Recewed
25 March
1985
PELLOW. S . L J HERBERG AND S E FILE Antagonrw~ of rhr cqjcc t\ oj rhc LII\~ICtrl hen~odicrzeprrrf, Ro 5-4864 on rrr[ruc rcr~rrul \clf-,/rrrrr,lofrotl,n /hc~rut PHARMACOL BIOCHEM BEHAV 24(Z) 193-197, 1986 -The effects of Ro 5-4864 (chlordlazepam) were exammed on respondmg for self-stunulation of the mid-lateral hypothalamus Rewardmg stimuli were delivered accordmg to a IO-set varlable Interval schedule of remforcement Ro S-4864 (l&30 mg/kg, subconsulslve dotes m these rats) decreased responding Thts effect was antagomzed by chlordtazepoxtde (5-10 mg/kg) and phenobarbltone (35 mg/kg) but not by the benzodlazepme receptor antagomst Ro 15-1788 (l&20 mg/kg), the hgand for penpheral benzodlazepme receptors PK I1 195 (60 mg/kg) or phenytom (60 mgikg) The pattern of mteractlons of Ro 5-4864 with these compounds differs from the pattern obtatned with other procedures, and suggests that Ro 5-4864 has effects on systems unrelated to anxiety, convulsive activity or sedatton Intracramal
self-stlmulatlon
Ro S-4864
Benzodlazepme
Ro 5-4864 15 a 1,4-benzodlazepme that, although havmg neghglble affmlty for ‘classical’ CNS benzodlazepme receptors, bmds with high affmlty to ‘peripheral-type’ benzodlazepme bmdmg sites that are present m the periphery [4] and m the CNS 1271 Although Ro 5-4864 has no benzodlazepme-hke actlvlty m ammal tests [31] and thus has long been consldered mactlve [24], recent mvestlgatlons mto Its behavloural effects have shown that It has convulsant [9, 22, 25, 301, anxlogemc [S. I I, 12, 191 and sedative [lo] propertles Further behavloural. electrophyslologlcal and blochemlcal mvestlgatlons have questioned the conclusion, made on the basis of bmdmg studies, that Ro 5-4864 does not Interact with the GABAlbenzodlazepmellonophore complex m the CNS, and suggest a possible action at the plcrotoxmm site that IS linked with the chlonde lonophore on this complex (see [ 19,201 for revtews) Respondmg for lateral hypothalamic self-stlmulatlon on a vanable Interval schedule of remforcement has been shown to be a sensltlve procedure for ldentlfymg and dlstmgmshmg the behavloural effects of benzodlazepmes, GABA agonists and antagonists, non-GABAerglc antlconvulsants [ 131 and benzodlazepme ‘antagomsts such as Ro 15-1788, CGS 8216 and FG 7142 [23,24] Our aim was to charactenze further the behavloural effects of Ro 5-4864 usmg this procedure, and to examme Its profile m combmatlon with several other compounds with which It has previously been shown to interact These
were
PK
I 119.5, an tsoqutnolme
‘Correspondence at the followmg address boulevard de I’HBpltal. 75634 Pans CCdex
carboxamtde Department 13, France
antagonism
tlve that, hke Ro 5-4864, selectively bmds to penpheral-type benzodlazepme bmdmg sites [ 15,161 and 1s thought to be an antagonist there [ 171, chlordlazepoxlde, a conventional I ,4 benzodlazepme with moderate potency for classlcal CNS benzodlazepme receptors but with neghglble affimty for penpheral-type sites [27], Ro 15-1788, an lmldazodlazepme that antagonizes several of the effects of benzodlazepmes and p-carbolmes [ 14,211 and that IS beheved to interact only with classlcal CNS benzodlazepme receptors [26], phenobarbltone, a barbiturate that displaces [‘HI-adlhydroplcrotoxmm and [ %S]-t-butyl blcyclophosphorothlonate from the plcrotoxm site, and phenytom, which Interacts with benzodlazepmes at low affinity bmdmg sites m the brain [2, 3,7, 18, 281 Doses of chlordlazepoxlde, Ro 15-1788, phenobarbltone and phenytom were chosen on the basis of previous studies ([13,23], Pellow and File, unpubhshed), dose-response data were determmed for Ro 5-4864 and PK I I I95 METHOD AnlnlUlJ
Male hooded PVG rats (Bantm and Kmgman Ltd ) weighing 230-250 g at time of surgery were mdlvldually housed with free access to food and water Under halothane anesthesia, twisted bipolar stainless steel electrodes (0 25 mm diameter, Plastic Products Inc ,
dertvade Pharmacologle.
193
Faculte
de Medecme,
Pltl&SalpCtniire,
Umverslte
Pan\
VI. 91
194
PELLOW, HERBERG AND FILE TABLE 1 MEAN ( + S E M ) S E L F - S T I M U L A T I O N R A T E (% B A S E L I N E ) IN E A C H 15-MIN PERIOD OF A 60-MIN TEST SESSION, IN RATS TESTED I M M E D I A T E L Y A F T E R IP INJECTION WITH Ro 5-4864 (10 AND 30 mg/kg) OR PK 11195 ( 1 0 ~ 0 mg/kg)
Drug
1st 15-mm
2nd 15-mm
3rd 15-mm
4th 15-mm
control Rol0 Ro30
982 _+ 1 03 952_+251 728_+ 6 16"
998 _+ 1 67 9 2 6 _+ 377 68 1 -+ 656"
975 _+ 3 61 952_+ 455 589_+ 104"
101 3 _+ 4 17 995_+ 503 634_+ 108"
PK 10 PK 30 PK60
952_+ 443 928 + 3 75 874-+ 723
997_+ 459 91 8 -+ 13 4 894_+ 750
101 5_+ 697 984 + 3 43 869_+ 800
962_+ 10 10 1006-+ 808 955 _+ 12 20
*p<0 05
R o a n o k e , VA) were stereotaxlcally implanted in the midlateral hypothalamus 1 0 m m posterior to Bregma, 1 3 mm to the left of the m,dllne and 8 0 m m below the skull surface E l e c t r o d e placements w e r e verified on enlarged photographic projects of unstained 50-/am frozen sections at the end of the investigation
150 i
125
"6 P
100
CONTROL
Ro 5-4864 (301
"/5
Drug~ Ro 5-4864 ( H o f f m a n - L a Roche), P K 11195 (Pharmuka), p h e n y t o m (Sigma), sodium p h e n o b a r b l t o n e ( B D H Chemicals) and Ro 15-1788 ( H o f f m a n - L a Roche) were suspended m distilled water with a drop of T w e e n 20 and dispersed by ultrasound, c h l o r d l a z e p o x l d e h y d r o c h l o n d e (Roche Products) was dissolved in distilled water Control injections consisted of dlstdled water, with or without a drop of T w e e n 20, as appropriate Drugs were injected ,ntrapentoneally ,n c o n c e n t r a t i o n s to give an rejection v o l u m e of 2 ml/kg
SelJ-Sttmulatton Rats were allowed to r e c o v e r from surgery for a week, and then trained to operate a pedal for a 0 5-sec 50-Ha s m e w a v e r e m f o r c m g impulse available on a variable interval schedule o f 10-sec m e a n duration (see [13, 23, 24] The stimulating current for each rat was fixed at the lowest intensity that elicited uninterrupted responding in preliminary trials in which intensities were d e c r e m e n t e d m decdog steps, b e t w e e n 50 and 10/xA r m s R e s p o n s e rates in different rats ranged b e t w e e n 15 and 25 l e v e r - p r e s s e s / m m During test sessions, animals w e r e allowed to selfstimulate for 45 mm, of which the last 30 mln provided a pre-mjectlon basehne The rat was then rejected, and allowed to respond for a further 75 mln If the rat stopped responding it was encouraged to restart by taps on the Fever, if this failed, by admlmstratlon of priming shocks, and, finally, by the e x p e r i m e n t e r placing the rat bodily on the lever R e s p o n s e rates w e r e r e c o r d e d automatically at 5-mm intervals as a dlg,tal printout Drug effects w e r e determined from the rate recorded during each 15-mln period following mjecUon, e x p r e s s e d as a percentage of pre-mjection basehne, and c o m p a r e d with the corresponding rate after control injections P/o( edut e
Expertment I Dose response data were obtained for Ro 5-4864 and PK 11195 on self-stimulation responding Drug
5O
_e 25
~,
o
Control PK CDP Pheny- Control PK CDP Pheny11195 (.5) O0) tom 11195 (5) (lO} tom (60) (60) (60} (60)
FIG 1 Mean (+S E M ) self-stlmulatzon rate (variable interval l0 sec) m the second 15-ram period after injection for rats given phenytom (60 rng/kg) or chlord,azepoxide (5 or 10 mg/kg) alone or m cornbmatlon with Ro 5-4864 (30 mg/kg)
groups were vehicle, Ro 5-4864 10 and 30 mg/kg, PK I 1195 10, 30 and 60 mg/kg ( n = 7 per group) L~pertment 2 The effects of an active dose of Ro 5-4864 (30 mg/kg) were investigated in combination wlth p h e n y t o m (60 mg/kg), p h e n o b a r b , t o n e (35 mg/kg), Ro 15-1788 (10 and 20 mg/kg), chlordlazepoMde (5 and 10 mg/kg), PK 11195 (60 mg/kg) ( n = 6 per group) Each rat was tested at 48-hr intervals, and m each exper,ment the drug treatments were administered m a predetermined random o r d e r that was different for each rat Statl~tt~ s
Data from E x p e r i m e n t 1 were analysed by Friedman 2-way analysis of variance, and c o m p a r i s o n s between individual groups were made with W d c o x o n matched-paws signed-ranks tests Data from E x p e r i m e n t 2 were analysed by W d c o x o n matched-pairs signed-ranks tests RESULTS
E~permwnt I Ro 5-4864 (10-30 mg/kg) caused a slgmficant dose-related reduction m responding for self-stimulation m the first, X-'=6 0, p < 0 05, second, X-'= 12 2 9 , p < 0 005, third, X-'=8 86, p < 0 05, and fourth, X-'= 10 57, p < 0 005, 15-mm time periods after injection (see Table 1) W d c o x o n tests showed that at 30 mg/kg slgmficant reductions m self-sumulatlon respond-
A N T A G O N I S M O F E F F E C T S OF Ro 5-4864
195 TABLE 2
S U M M A R Y T A B L E OF T H E ABILITY O F S E V E R A L C O M P O U N D S TO A N T A G O N I S E T H E E F F E C T S O F Ro 5-4864 IN A N I M A L TESTS O F ANXIETY, C O N V U L S I V E ACTIVITY, E X P L O R A T I O N A N D S E D A T I O N , A N D IN T H E P R E S E N T S E L F - S T I M U L A T I O N P R O C E D U R E
Drug Benzodmzepmes Barbiturates Ro 15-1788 PK 11195 Phenytom
Anxiety ~ [111 ~* - [Ill - [8,12] ~ [8]
Convulsions
Exploration
~ [7,29] ~ [291 ~ [71 $ [6,8] ~ [8]
Motor
~ ~ - [10l $ [10] - [10]
') ') - [101 - [10] [10]
ICSS
-
*Pellow and Fde, unpubhshed data compound antagomses Ro 5-4864, - compound does not affect Ro 5-4864, '~ not yet reported References to studies are in brackets (see text)
mg were observed at all time-points PK 11195 had no s)gmf)cant effect at any t~me period, X-'=2 49, 2 48, 0 09, 0 43, respectwely), see Table 1
150
CONTROL
Ro 5-4864 (30)
lz5
I~ ~ p e r t m e n t 2
"5 100
Phenytom (60 mg/kg) s)gmficantly decreased selfst~mulat)on m the second 15-mm period after mjecUon (p<0 05) but d)d not antagonize the reduction produced by Ro 5-4864 (30 mg/kg) (see F~g 1) CDP increased selfst)mulat)on at 5 mg/kg (p<0 05) and decreased )t at 10 mg/kg (p<0 05), and at both doses s~gmficantly reversed the reduct)on produced by Ro 5-4864 (30 mg/kg, p < 0 05, see F~g 1) Interestingly, however, Ro 5-4864 d~d not antagomze the mcrease m self-stimulation produced by CDP (5 mg/kg) PK 11195 (60 mg/kg) had no s)gmficant effect on self-st)mulaUon alone, and d~d not antagomze the effects of Ro 5-4864 (see F)g 2) Ro 15-1788 (10-20 mg/kg) d)d not antagomze the effects of Ro 5-4864, although at 20 mg/kg th)s compound when g)ven alone s)gmficantly increased self-sUmulaUon (see F)g 2) Phenobarb)tone (35 mg/kg) s)gmficantly increased self-st~mulat)on (p<0 05), and also antagomzed the reduced self-sUmulaUon produced by Ro 5-4864 (30 mg/kg, p < 0 05, see F~g 2) DISCUSSION
Ro 5-4864 at 30 mg/kg produced a decrease m responding for lateral hypothalam)c self-st~mulauon, s,mflar to that reported for the benzodmzepme receptor 'reverse agomsts' CGS 8216 and FG 7142 [23.24] Since Ro 5-4864 is a compound w)th known convulsant acuons, ,t ~s necessary to cons)der the extent to wh)ch these act)ons could account for )ts effects on self-stimulation The dose of Ro 5-4864 that has been found to effect,rely cause convulsions has vaned conslderably, both between (c f [1,6, 25, 29]) and within (cf [6, 7, 22]) laboratories In our expenence, d)fferences have been observed between species (cf. [6,7]) and stratus (cf [6,9]), and also between different batches of the supphed compound In each new batch of experiments, then. we have found ,t necessary to estabhsh the effective convulsant dose of Ro 5-4864 before carrying out the intended behavloural work During the course of the present study we observed, m th,s strain of rats, that convulsions did not occur at doses below 90 mg/kg At 90 mg/kg 100% rats convulsed, at 60 mg/kg we only observed myoclomc spasms m one rat who had been treated and tested for self-st~mulaUon responding In no cases were convulsant-hke activities observed at 30
75 -5
Control
15-1788 Pheno (10) (20) barb (35)
Control
15-1788 Pheno(10) (20) barb (35)
FIG 2 Mean (=S E M ) self-sumulatJon rate (variable interval, 10 sec) m the second 15-mm period after rejection for rats given PK 11195 (60 mg/kg) and Ro 15-1788 (10 or 20 mg/kg) alone or m comblnaUon with Ro 5-4864 (30 mg/kg)
mg/kg It is therefore unlikely that the convulsant properties of Ro 5-4864 were contributing to ~ts effects on selfst~mulauon The pattern of mteracUons of Ro 5-4864 w~th other drugs (see below and Table 2) supports this view PK 11195 alone (10-60 mg/kg) had no stgmficant effect on self-st~mulaUon, htgher doses were not investigated since at the 60 mg/kg dose some ammals showed unusual body postures that could have interfered w~th their ablhty to lever-press The reducUon m self-stimulation produced by Ro 5-4864 (30 mg/kg) was not antagomzed by PK 11195 (60 mg/kg), this is consmtent with the inability of PK 11195 to antagomze the effects of Ro 5-4864 m the socml Interaction and pumshed drmkmg procedures [12]. although PK 11195 does antagomze the proconvulsant and convulsant effects [1,9] and the reductions m exploratory head-dipping produced by Ro 5-4864 [10] As found previously [13, 23, 24], chlordlazepoxlde (5 but not 10 mg/kg) Increased self-stimulation Chlordmzepoxlde at both doses sJgmficantly reversed the reducuon m response rates produced by Ro 5-4864, however, interestingly, Ro 5-4864 was unable to antagomze the increase m selfstimulation produced by chlordmzepoxlde (5 mg/kg) Chlordmzepox~de is able to antagomze the anxlogemc effects of Ro 5-4864 [11], and other benzodmzepmes have been found to antagomze the convulsant effects of th~s compound [7,29] Th~s )s s~mdar to the effects obtained by Herberg and Wllhams [13] w)th plcrotoxm and chlordmzepoxlde, however, unhke plcrotoxm. Ro 5-4864 did not produce a further
196 e n h a n c e m e n t o f r e s p o n d i n g w h e n c o m b i n e d with chlord l a z e p o x l d e (10 mg/kg) R o 15-1788 (20 mg/kg) e n h a n c e d self-stimulation, as previously f o u n d b y P e | l o w et a! [23] This c o m p o u n d did n o t a n t a g o n i z e t h e r e d u c t i o n in s e l f - s t i m u l a t i o n p r o d u c e d b y Ro 5-4864, again c o n s i s t e n t with findings in t h e social i n t e r a c tion t e s t o f a n x i e t y a n d the h o l e b o a r d [10,11] H o w e v e r , R o 15-1788 (20 mg/kg) IS able to a n t a g o n i z e the c o n v u l s i o n s p r o d u c e d b y R o 5-4864 in at least s o m e c i r c u m s t a n c e s [7] P h e n o b a r b l t o n e (35 mg/kg), like the b a r b i t u r a t e p h e n o b a r b l t o n e [13], i n c r e a s e d s e l f - s t i m u l a t i o n a n d a n t a g o n i z e d t h e effects o f Ro 5-4864 Again, similar r e s u l t s h a v e b e e n obt a i n e d m t h e social i n t e r a c t i o n test (Pellow a n d File, u n p u b h s h e d ) , w h e r e a d o s e o f p h e n o b a r b l t o n e t h a t also i n c r e a s e d social m t e r a c t i o n w a s able to r e v e r s e the d e c r e a s e p r o d u c e d b y R o 5-4864 H o w e v e r , it is n o t p o s s i b l e to be sure w h e t h e r t h e a n t a g o n i s m b e t w e e n t h e s e t w o c o m p o u n d s in b o t h of t h e s e t e s t s is specific o r w h e t h e r it simply reflects a cancell a t i o n o f t w o o p p o s i t e intrinsic a c t i o n s B a r b i t u r a t e s also p r e v e n t the c o n v u l s i o n s usually o b s e r v e d with Ro 5-4864 [29] C o n s i s t e n t w i t h t h e r e p o r t of H e r b e r g a n d Williams [ 13], p h e n y t o l n d e c r e a s e d s e l f - s t i m u l a t i o n T h e Inability of p h e n y toin (60 mg/kg) to a n t a g o n i z e the d e c r e a s e in s e l f - s t i m u l a t i o n p r o d u c e d b y R o 5-4864 c o n t r a s t s with its ability to antagonize the a n x l o g e n l c a n d c o n v u l s a n t a c t i o n s o f this c o m p o u n d [8,9] b u t IS similar to its inability to a n t a g o n i z e t h e r e d u c h o n s
PELLOW, HERBERG AND FILE in e x p l o r a t o r y head-d~pplng a n d l o c o m o t o r activity in the h o l e b o a r d p r o d u c e d b y R o 5-4864 [10] It ~s unlikely t h a t the d e p r e s s a n t effect that p h e n y t o i n p r o d u c e s w h e n given a l o n e w o u l d interfere with its ability to a n t a g o n i z e the r e d u c t i o n p r o d u c e d by Ro 5-4864, since c h i o r d m z e p o x l d e at a d o s e t h a t d e c r e a s e d r e s p o n s e r a t e s (10 mg/kg) was stdl able to antagonize the r e d u c t i o n p r o d u c e d b y Ro 5-4864 In c o n c l u s i o n , the p a t t e r n of i n t e r a c t i o n of Ro 5-4864 with o t h e r c o m p o u n d s m the v a r i a b l e - i n t e r v a l self-stimulation p r o c e d u r e s h o w s a b r o a d similarity to the p a t t e r n s s e e n in v a r i o u s o t h e r p r o c e d u r e s , b u t t h e r e are also s o m e n o t a b l e d i f f e r e n c e s (see T a b l e 2) It ~s t h u s u n c e r t a i n w h e t h e r the effects o f Ro 5-4864 in this p r o c e d u r e are to be a t t r i b u t e d to its effects o n a n x i e t y , c o n v u l s a n t activity or s e d a t i o n , or to ~ts effects on o t h e r , u n r e l a t e d s y s t e m s T h e s a m e q u e s t i o n applies to s e v e r a l related a g e n t s t h a t h a v e also b e e n f o u n d to d e p r e s s s e l f - s t i m u l a t i o n , s u c h as the b e n z o d m z e p m e recept o r ' i n v e r s e a g o n l s t s ' F G 7142 a n d C G S 8216, a n d the recept o r a n t a g o n i s t Ro 15-1788 [23.24] E x p e r i m e n t s are c u r r e n t l y u n d e r w a y to i n v e s t i g a t e th~s q u e s t i o n
ACKNOWLEDGEMENTS SEF ~s a Wellcome Trust Semor Lecturer We are grateful to the Medical Research Councd for eqmpment
REFERENCES 1 Benawdes, J , F Gullloux, D E Allam, A Uzan, J Mlzoule, 13 Herberg, L J and S F Wdhams Ant~confl~ct and depressant effects by GABA agonists and antagomsts, benzodlazepmes and C Renault, M C Dubroeucq, C Gueremy and G Le Fur non-GABAerglc antlconvulsants on self-stimulation and locoOpposite effects of an agomst, Ro 5-4864, and an antagomst, PK motor activity Pharma~ol Btoc hem Behm 19: 625-633, 1983 11195, of the peripheral type benzodmzepme binding sites on 14 Hunkeler, W , H Mohler, L Plen. P Polc, E P Bonettl R audlogemc seizures m DBA/2J mice Life S~; 34" 2613-2620, Cumin, R Schaffner and W Haefely Selective antagonists of 1984 benzodlazeplnes Nature 290: 514-516, 1981 2 Bowhng, A C and R J DeLorenzo Micromolar affimty ben15 LeFur, G , F Gudloux, P Rufat, J Benavldes, A Uzan, C zodlazeplne receptors identification and characterlsatlon m Renault, M C Dubroeucq and C Gueremy Peripheral benzocentral nervous system St wnce 216" 1247-1249, 1982 dlazeplne binding s~tes effect of PK 11195, I-(2-chlorophenyl)3 Bowhng, A C and R J DeLorenzo Multiple phenytom bradN-methyl-N-(I-methylpropyl)-3-1soqumohne carboxamlde II In mg sites ldentlficatmn and charactensaUon m brain membrane vlvo studies Lth" 3~t 32. 1849-1856, 1983 So~ Neuros~t Abstr 8. 505, 1982 16 LeFur, G , M L Pemer, N Vaucher, F Imbault, A Flamler, 4 Braestrup, C and R F Sqmres Specific benzodmzepme recepJ Benavldes. A Uzan, C Renault, M C Dubroeucq and C tors m rat brain characterized by high-affinity [3H]-dmzepam Gueremy Peripheral benzodlazeptne binding sites effect of PK binding Pro~ Natl Acad S~t USA 74: 3805-3809, 1977 11195, l-(2-chlorophenyl)-N-methyl-N-( l-methylpropyl)-35 Burnham, W M , L Spero, M M Okazakland B K Madras isoqumollne carboxamlde I In wtro studies Lt/e St; 32 1839Saturable binding of [3H]-phenytom to rat brain membrane 1847. 1983 fractmns Can J Physlol Pharma~ ol 59. 402-408, 1981 17 LeFur, G . N Vaucher. M L Perner, A Flamler, J Ben6 File, S E Pro- and ant~-convulsant properties of PK 11195, a awdes, C Renault, M C Dubroeucq C Gueremy and A hgand for benzodmzepme binding sites development of Uzan Differentiation between two hgands for peripheral bentolerance Br J Pharmacol 83: 471-476, 1984 zodlazeplne binding sites, [ ~H]-Ro 5-4864 and [ ~H]-PK I 1195, by 7 Fde, S E , A R Green, D J N u t t a n d N D Vincent On the thermodynamic studies Life S~t 33: 44%457, 1983 convulsant actmn of Ro 5-4864 and the existence of a m~cromo18 Okazakl, M M , B K Madras, K E Llwngston, L Speroand lar benzodmzepme binding s~te m rat brain Psv~ hopharma~ olW M Burnham Enhancement of [~H}-phenytom binding by ogv (Berhn) 82: 19%202, 1984 dmzepam and bJcuculhne Life S~I 33. 409-414, 1983 8 File, S E and R G L~ster The anxlogemc action of Ro 5-4864 19 Pellow, S and S E Fde Characteristics of an atypical benis reversed by phenytom Neuros~t Lett 35" 93-96, 1983 zodmzepme, Ro 5-4864 Neuro~l Bu~behm Re~ 8 405-413, 9 File, S E and P S Mabbutt Behav~oural effects of Ro 5-4864, 1984 a hgand for the m~cromolar benzodmzepme receptor Br J 20 Pellow. S and S E File Behawoural effects of Ro 5-4864 a Pharma~ol 78: 76P, 1983 peripheral-type benzodmzepme "~Life St t 35" 22%240, 1984 10 Fde, S E and S Pellow Ro 5-4864, a hgand for benzodmzepme 21 Pellow, S and S E Fde Multiple sites of action for anxtogemc m~cromolar and peripheral binding sRes antagonism and drugs behawoural, electrophysmloglcal and biochemical correenhancement of behavmural effects Psy~hopharrna~ ology lations Psw hopharma~ olog~ (Berhn) 83:304-315, 1984 (Berhn) 80: 166-170, 1983 22 Pellow, S and S E Fde Proconvulsant and antlconvulsant 11 File, S E and S Pellow The anxmgemc actmn of Ro 5-4864 drug effects m combination w~th the convulsant benzodmzepme effect of chlordlazepox~de, Ro 15-1788 and CGS 8216 Naunvn Ro 5-4864 I PhaHn Pharma(ol 37" 560-563, 1985 S~hmwdebergs Arc h Pharma~ ol 328: 225-228, 1985 23 Pellow, S , S E File and L J Herberg lntracranml self12 File, S E and S Pellow The effects of PK 11195, a hgand for stimulation d~stlngmshes between two benzodlazepme benzodmzeplne binding sites, in ammal tests of anxiety and antagomsts Neuros~t Lett 47" 173-177. 1984 stress Pharmat ol Bto~hem Behav, in press, 1985
A N T A G O N I S M O F E F F E C T S O F Ro 5-4864
24 Pellow, S , L J Herberg and S E Fde The effects of the /3-carbohne, FG 7142, on mtracranml self-stimulation in the rat Pharmac ol Bto~ hem Behav 21: 667-669. 1984 25 Plen, L , P Polc, E P BonettL W Burkard, R Cumin, R Scherschhcht and W Haefely Some pharmacologxcal effects of Ro 5-4864, a specific hgand for the peripheral type of benzodlazepme binding sites N a u n v n S( hmtedeberg~ Arc h Pharma
197
28 Shah, D S , P Chambon and A Guldottl Binding of[~H]-5,5 dlphenylhydantoln to rat brain membranes Neuropharma~ ologv 20. 1115-1119 1981 29 Welssman, B A , J Cott, D Hommer, S Paul and P Skolmck Electrophyslologlcal and pharmacological actions of the convulsant benzodlazepme Ro 5-4864 Lttr I Pharma~ ol 97. 257263 1984 30 Welssman, B A , J Cott, S M Paul and P Skolmck Ro 5-4864 a potent benzodlazepme convulsant Lur I Pharma~ ol 90. 149-150, 1983 31 Zbmden, G and L O Randall Pharmacology of benzodlazepmes laboratory and chmcal correlations Ad~ Pharmayo/5 213-291, 1967