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Evaluation of (d -Pro2, d -Trp7,9)-substance P as an antagonist of substance P responses in the rat central nervous system
Nentroscience Letters, 30 (!982) 291-295 Elsevier/North-Holland Scienlific Publis!~rs L:d.
291
EVALUATION OF (D-Pro ~, D-TrpT,~).SUJSTANCE P AS AN AN2AGONIST O]F SUBSTANCE P RESPONSES IN T~.~.ERAT CENTRAL NERVOUS SYSTEM
T.E. SAI.T, G.J. DE VRIES*, R,E. ROOR1GUEZ *®, P.M.B. CAH.I.ISAC, k. M O g R I S and R,G. H I L L
Department qf Pharmacology, University of Bristol, Med,ical School, Bristol (U.K./ (Received April Ist, 1982; Accepted April 7th, 1982)
The (D-Pro:, o-'l-rp7,9) analogue of substance P has Vecn tested ;'or subslar~e .o antagonist ac!~,tty ~n the catdal trigeminal nucleus in ~ivo, an',:l in the isolated spinal cord in vitro. In neither tale ;-'-.~, tnc anaI,ogue found to be a specific antagonist of substaitc¢ ~, although the analogue did have v,'~'ak amagorfist actions in the isolated guinea-pig ikum preFarati~,-~. It is concluded ~N~t the ana!ogue is nut a suitable tool for the identification of putative subst~:nce P systems in tee spine! cord or the caudM trigeminai nucleus of the rat.
There is evidence that the undecapcptidc substance P (SP) nzay bc a neurotransmitter in various parts of the central nervous s y s t e m , b'.-~t i.~¢egti~'~tio~.~ ~. have l:een hampered by the lack of st~ilable ~.ntago~isls f,~r ~n!s .t~.:':id:--[II]. However, synlhetic analogues of SP wi,h aiitagonist propcrti':s ha¢¢ b..'~'n n:cc~lily developed 13i. In particular, (rePro 2, ~s-Trp'~,9)..SP{DPDT-SP) has be,.m showr, to bc a speed it SP a~,tagonist on smooth muscle preparations [i0], and to "~.'.~.',,.,o¢.'iz," it-.:,. action. 9f SP on neurones in tl'~ locus coe.ru|eu:, [2j. A~, SP may hc invol'~.~! h: t~tst:~.-~ transmission ie the spinal cord and caudal trigemina! nucleus (n. c~udr, IL.',) !6~, it :~ imporlant to ehtablish whether or not DPDT-SP is ~ suitable tool i~or the stttdv of putative SP pathways at these sites. We have therefore s;tudicd the effects of DPDTSP on respons,:s, to SP of single neurones in n ¢aud?,lis, a restral cxterislon 3f the dorsal horn of the spinal c~rd [4|, and or, SP-induced depolarizations ,of motoneurones recorded from ~he ventral roots of the isolated spinal cord. Extraeeltular recordings were made from single n. czud~!is neurc.ne~ ha r,ta[c, urethane-anaesthetized, Wistat rats ( 4 ~ 0 - 5 ~ g',, using the 4 M NaCi-fi[led ~:~:lt~'e barrel of multibarr~Ied micropipettes, hhc outer ~,arre~s of which were ,.tsed for :he iontophoretic a:~p~ication of drugs (Yig, l) [I"41. SP was f~un'3 to have '..'x~:itau~ry
* P~rmar~.,',nt address: .~:et.h~rtands. l'.'t._,.,,.,,,r- f-~r r~,,.-~,,., }t.ee~:,'v-h. ", ..r, .... ;~-,,...., ,"8<:':., ~ ~.. I f:.Cr9 ~,is... .' t ' ,",~::~,, "".. Th': Netherlands. | "F " " Permal~nt ~lort. ~:v,: C'oi::gic de M.,~ai.:ira c,: .'wila, Uni:'cr:A:y ..>~";'=i:.,,~,e.,;.~. ~ii,;.;;:.
0304-394,0182/',ggdO-£-~)/$ 02.75 ~.. t9~2 Et.g~:vi,.'r,'North-iT )i'..arld !~cier,:if~c Publi~h.-.r'g i.~d.
292
,
! .......
.
.
.
.
.
.
.
.
k--G~a5 t,..A
i
~-':~ 50]
II ,] = = I I I I
OPOT-SPP$ ~ = I I l11
I IL-r..fl.u$Ot~
O] Fig. I. Analogue ratemeter tract from a n. caudalis neurone excited by SP (40 nA, ~'-2)in the present t of t -glutamate (5 nA.~. Drug~ w,:re a0pli¢~l from barr¢t~ :ontaining either sodium t-ttlutama:e (Sigma, 0.5 M, r~H 8.5), S." (i~enirtsula, 2.3 mM in 150raM NaCI) or EJPi)r-sP (Peninsu|a, 3 r a m in tSOmM NaCI, or 3 mM in ?.f)mM sodium acetate, p;-t 4.5). In addition, one barrel of the micropipette contained " M Nat?l for current balancing and current controls, and am-,thtr barrel Pontamine sky blue dye (2,5% in acetate bqffer) for histological verification o~ recording ilr s. DPDT-SP t45 hA, ~ ) was found to dcpre~r, SP respon,~cs (upper trace), t:ut v, hen tested agaiast j °glutarnate.a.voktxt e~citation.,, ( I , lower ~racc), the analogue ',~as found to de!~re',,, tt~esc also.
actic.n,~ when ejected (20--80 nA) onto neurones with a pre-existing ongoing firing .,'ate (eill~er Sl:ontaneous or t-glutamate-evoked), as previously described [13, 15]. When challenged with concurrent applicalion of DPDT-SP (30-150 nA), respons:s to SP were reduced in 6 of 17 neurones (lqg. i). However, in all of these ca.',e,i there v,a:~ also a marked reduction of the ongoing firing rate of th., neurone and/,)r depression of the L-glutamate-evoked activity (Fig. l). Tlr~,e effects of DPDT-SP thus cannot be regarded as examples of specific SP antagonism. On the remaining 11 ncmrones, no depression of SP respon~:s ~ as evident, and the analogue either had no effect (n = 6) or had excitatory actions (n = 5). Excitatory effects tyl~cally consisted of increases in firing rate and paroxysmal 0ursts of firing, which were often ~'ollowed by depression of the neurone. In isolated spinal cord preparations, SP (0. l-2.5 #M) was found to depolarize motoneurones, as previously demribed [9] (Fig. 2). When DPDT-SP was su~rfused at similar concentrations, no pronounced ¢~irect effects were noted, and SP responses in the presence of the analogue were. eithcr similar to or slightly greater than cor, trol responses. When higher concentrations of DPDT-SP were applied (up to 20 #M), direct debolarization of the motoneurones became evident, and SP reslaons,"s :,,ere generally potentiated (Fig. 2). in order to enable a comparison to be. made, between the central and lx'ripheral actions of DPEr'F-SP, the analogue was also tested for SP antagonist action on four
293
frill/i "
SP 7..5,t~W
il
I
r--1 DPDT-SP20~.,t."
~t
[
P-~T-
1
SP
ImV
]
B~M
,.
"
~)mir~
Fig. 2. 0(2 rec~dings o f motoneurone polarization were made from the lumbar ventral roots of .1 hemisected isolated spinal ~ r d preparations taken from 4-g-day-old tats, and drugs we:e appJied by additiort t o the bathing medium [121. The figure shows depolarization~ ~roduced by 2.5 ~M SP superfusion ( I ) before, during and after superfusion with DPDT-SP (~--.21,8 It.M). The analotuc potentiates SP responses, and when applied at 20 uM (D) produces a direct depolarization.
A
--
i DiI~T-SP o
o
i011
o
o
oSP
~----o
'+J+1
//t /
o / / /
/
//(
i,t
/
/
a--~ SP ¢~lone
/
/ +/
0J It
Fig. 3. A: contraction.~ of a guinea pig i!eum to addition of SP (% 2.5 × I0 ~)M ) to the bathin:; mcdiun:. Addition of DPDT-SP (e, I 0 ~' M) al~ o prodttces a contraction, following which re:+ponszs to sir, are con.qd-.rahty redttced. Breaks in the record corr,'s2.~3".d to periods of washing of the ti+,~tte. B: do~--respons¢ curves of I|,e same ileur as in (A) to SP in the r.resence (*) anti abscn:e (31 ¢,f I h " M DPDT-SP. It ,.'an be seen thai the anaiot, ue shifts ~.he SP :esponse curve to the right, and it i,. s,.ll po:,siblc to achieve a roa'~imal SP response in lhe presence or DPD'I-SP.
guinea-pig ileum preparations 11, 31. SP (10 ...`7 M - 5 × :,0 '* M) was found to pro:tuce contractions of the ileum. Addition of DPDT-SP to tt~e batbh~r., medium ( I 0 - ; M - - I O t, M ) was also fc~;nd .to pro~uc,: c.:).;.~r;.,c::..~;n,; :;n+, :,f:e: ~hc.,:c had :mbsided, responses to SP wet'(, reduced (Fig. 3A), as has been described !.~ od~cr ' . swooth muscle preparalions [|0] . The redaction (,f ""~[- ~c"4;,.~:~... Ul;.~,~..~.~~. .'..~b,- ( ;"J / ; to competiliv~ antagonism rathe~ than densensitizati,:~.n, as rna.'.-.:ir:,z:[ ,.:o~+.~r-.cti,.m:,
29~
could still be ~,chieved in the presence of the antagonist by increasing the 5P dose, and the dose-response curve for $P was foead to be shifted to the right by DPDTSP (Fig. 3B), as previously de-.~m"ae~.,[ 1 0 ] , , . . . . , : . . . . SP-induced excitations oi locus ¢o¢~1¢us ~ [ ~ ] . H .o~evet~.em" f i ~ : h ~ < , a . cauda|is and the spinal cord suggest that lids aaaloguedoes ao¢ l m v ¢ ~ ~ properties, or that if it has, that t~cs¢ properties ate masked by other effec'.s of DPDT-SP that appear at concentrations insufficient to produce antagonism. The finding that considerably g:cater amounts of the analogue than of SP are required to produce SP antagonism in smooth mL~cle preparations (Fig. 3) [10| would support this suggest.ion. A further implication of these :¢sults is that recel~OrS for SP in the locus ¢oeruleus may be 4ifferent to those in the spinal cord and n. caudalis, the former being ~:aore sensitive to the antagon:~st actions and less sensitive to the direct effects of DPDT-SP. [t is also noteworthy that DPDT-SP has been fcund to be have neurotoxin-like aclions [7], and it is po~.~sibte that these effects may be related to the direct effects seen in this study. in conclusion, it appears that DPDT-SP is not a suitable tool for the study of putative SP systems in the spinal cord or n. caudalis of the rat, although this cornpound has prov-d u~ful in the peripheral nervous system [10] and in other parts of the central nervous system [2]. Th..~e results provide further evidence for the exi.qt,:ncc of sub-tyl~s o" SP receptor o~ different sites in the body [5, :l]. Wc wi,,h to :hank Dr. R.H. Evan~ for as-,/stanc¢ with the spinal ~:o~:lpreparation. ~t:pr ;rtt:d by gran;:~ froo~ the M.R.C., Royal Society and S.R.C. t [:dml)t+rrb Stati, Phal':~a+<.>]o+Acal l:~;'+;..r;Inel~t.> on i+,ol++t,.xl Pre+'.~+ralion,,. 2l~c+ end., Ci)urch!ll. I.iving,+tc,+,, Edi:+burgh, t9"t+, pp. 2~-2~., 5B-66, 152--153. 2 En,, bcrp.. G., Svc,m,,~'m, T.H., Ro.~¢ll, S. and Felkers, K,+ A ',ynl~+.k: peptich~ as an amaloni+..t of ,~ub~tance P, Nature (I.on~i,L 29~ ~19~.!) 222-223. 3 I'~lk¢,s, K., H6rig, J., Rdsell, 5. and BjO~'ktoth. U.. Chemical desi):n of attlag¢)tli:,l~ t bUbSt&nCf P, ..\eta physic.!. ~cand., ! I ! (19gl, 505-.506. .~ G(,t)¢l, S., Fall~, W.M. and Hockfic~d, S., "l'h¢ divis~.oo of the d o t ~ ! and vei~tfal horn= of lh© mamtn~han caudal medulla inlo eight layers ush~$ anatomk-ai ~:r=;.,,-ria. In D. Andcru~rJ and 13. M.~lthcws (Ed~.), Pain ia =he i'rigemina~ Ro~ion, El~vicr/N,:v:a-H~,~u~d Biomedit~ Pt¢~, An:slc,d=t,~, 1977, ?p. 443-453. '~ }tawco,ck, A.B., Ita],'¢s, A.G. and Ty=.rs, M B.. Aggni,_: prop¢~i¢~, of (t~-P~.o-Phe'r,t~-TrTp *) gLt~',~t;ilZCC P ill vitro - c~i(1¢t~¢¢ t'o~ mote than on¢.ty[~ or substan¢-. P tc~,-~ot, Briz. J. Pharra~.¢.=l., 75 ~t982) l i ~P. 6 Henry. I.I : S~d~.,;:;wce P ~ad poin: an tlod;~ting, "lr~.~ld~, NcuraSci , 3 (l':Yd0) 9~-97. "' t-4(~kfch, | . , Viqc¢~t, S., ~tc~ls,ca, L., Ro~¢ti, S., }'~R,~:~s, K., Marr, cy~ K., Gold,tell1, ~.. ~=tv,J Ct'~c)lc. C.. tmm~i~t~h;.stt×'hemi,,~d ev!denc~ for a "r~*~rozoxk.,' a~;tio~ of (o-Pto2,o-Ttp;.~}-substltrtce P, ~:~ a::a!~,gu¢ ,.~i=h sut'stanc¢ P ~magonistic activity, Ac:a physiol, stand., 113 (198I) 571-573. g P,cr:,cn. L.L.: l.e¢, C.M., ,Land[~:~g, B,E.B. and Watson, S.P., Differins rank orders of ~ t e n c y of ~ubstan¢¢ P a ~alo~lt:¢~ .;u~.(:e,,: ih? ¢.xii!ertce o | multiple reccptct sub.l.vpe.~, P~ti:. J. Ph,tl'~a¢ol.. "/5 (!982) ~.12P.
295 9 Konis~, S. ~ Otzuka, M., Excitatory action of hypothatamic substance P on spinal mote,net, onrs of newborn rats, ~,latur¢ (Lond.). 252 (19"/4) ?34-735.
!0 Leander, $., l-ltlkmson, R., Resell, S., Folkers, K., Sundler, ). and Torp.qvist, K., A .~pccific sub~taace P ~mtalonist l:,k~ckssmooth muscle contractions in.duced by non-choii~.,cL~ic,nonadreneri~ here sfimutatk;n, Nature (Lond.), 294 (I~I) 467-469. 11 Nicoll, R~A,, Sct~.-nk~, C, and L.ceman, S.E., Substance P as a transmitt..'r candidate, Ann. Roy. bleurosci., 3 (1950~ 227-26& 12 Otsuka, M, and Konishi, S., Electrophysiology of mammalian spinal cord in "ilro, Nature (Lond.), 252 (!974) "I33-734. 13 Salt, T.E., Ber,y, 'JlC., Hill, R.G. and Morri:+, R., The effects of p~ptide neuro:ransmitter can~didates on single neuroncs in the rat trigcminal nucleus caudalis. In B. Matthews and R.G. Hill (F:ds.), Anatomical, Fhysiological and Pharmacological Aspects of Tri~cminal Pain, Elsc':icr/NorthHolland Biomcd!cal Press.. Am.~tcrdam, in press. 14 Salt, T.E. and Hill. R.G., Differentiation of excitatory amino acid r.'ccoTor, in h e rat ¢a:~dai trige:ninai nuclc:Js: a microiontophoretic study, Neuroph~rmacology, in prt s',. 15 Zieglgtlnsberger, W. and Tulloch, i.F., Effects of substance P on ncurot,es in the dot-;~~l horP. of th,? spinal cord of t,e cat, Brain Res., 166 (1979) 273-282.
291
EVALUATION OF (D-Pro ~, D-TrpT,~).SUJSTANCE P AS AN AN2AGONIST O]F SUBSTANCE P RESPONSES IN T~.~.ERAT CENTRAL NERVOUS SYSTEM
T.E. SAI.T, G.J. DE VRIES*, R,E. ROOR1GUEZ *®, P.M.B. CAH.I.ISAC, k. M O g R I S and R,G. H I L L
Department qf Pharmacology, University of Bristol, Med,ical School, Bristol (U.K./ (Received April Ist, 1982; Accepted April 7th, 1982)
The (D-Pro:, o-'l-rp7,9) analogue of substance P has Vecn tested ;'or subslar~e .o antagonist ac!~,tty ~n the catdal trigeminal nucleus in ~ivo, an',:l in the isolated spinal cord in vitro. In neither tale ;-'-.~, tnc anaI,ogue found to be a specific antagonist of substaitc¢ ~, although the analogue did have v,'~'ak amagorfist actions in the isolated guinea-pig ikum preFarati~,-~. It is concluded ~N~t the ana!ogue is nut a suitable tool for the identification of putative subst~:nce P systems in tee spine! cord or the caudM trigeminai nucleus of the rat.
There is evidence that the undecapcptidc substance P (SP) nzay bc a neurotransmitter in various parts of the central nervous s y s t e m , b'.-~t i.~¢egti~'~tio~.~ ~. have l:een hampered by the lack of st~ilable ~.ntago~isls f,~r ~n!s .t~.:':id:--[II]. However, synlhetic analogues of SP wi,h aiitagonist propcrti':s ha¢¢ b..'~'n n:cc~lily developed 13i. In particular, (rePro 2, ~s-Trp'~,9)..SP{DPDT-SP) has be,.m showr, to bc a speed it SP a~,tagonist on smooth muscle preparations [i0], and to "~.'.~.',,.,o¢.'iz," it-.:,. action. 9f SP on neurones in tl'~ locus coe.ru|eu:, [2j. A~, SP may hc invol'~.~! h: t~tst:~.-~ transmission ie the spinal cord and caudal trigemina! nucleus (n. c~udr, IL.',) !6~, it :~ imporlant to ehtablish whether or not DPDT-SP is ~ suitable tool i~or the stttdv of putative SP pathways at these sites. We have therefore s;tudicd the effects of DPDTSP on respons,:s, to SP of single neurones in n ¢aud?,lis, a restral cxterislon 3f the dorsal horn of the spinal c~rd [4|, and or, SP-induced depolarizations ,of motoneurones recorded from ~he ventral roots of the isolated spinal cord. Extraeeltular recordings were made from single n. czud~!is neurc.ne~ ha r,ta[c, urethane-anaesthetized, Wistat rats ( 4 ~ 0 - 5 ~ g',, using the 4 M NaCi-fi[led ~:~:lt~'e barrel of multibarr~Ied micropipettes, hhc outer ~,arre~s of which were ,.tsed for :he iontophoretic a:~p~ication of drugs (Yig, l) [I"41. SP was f~un'3 to have '..'x~:itau~ry
* P~rmar~.,',nt address: .~:et.h~rtands. l'.'t._,.,,.,,,r- f-~r r~,,.-~,,., }t.ee~:,'v-h. ", ..r, .... ;~-,,...., ,"8<:':., ~ ~.. I f:.Cr9 ~,is... .' t ' ,",~::~,, "".. Th': Netherlands. | "F " " Permal~nt ~lort. ~:v,: C'oi::gic de M.,~ai.:ira c,: .'wila, Uni:'cr:A:y ..>~";'=i:.,,~,e.,;.~. ~ii,;.;;:.
0304-394,0182/',ggdO-£-~)/$ 02.75 ~.. t9~2 Et.g~:vi,.'r,'North-iT )i'..arld !~cier,:if~c Publi~h.-.r'g i.~d.
292
,
! .......
.
.
.
.
.
.
.
.
k--G~a5 t,..A
i
~-':~ 50]
II ,] = = I I I I
OPOT-SPP$ ~ = I I l11
I IL-r..fl.u$Ot~
O] Fig. I. Analogue ratemeter tract from a n. caudalis neurone excited by SP (40 nA, ~'-2)in the present t of t -glutamate (5 nA.~. Drug~ w,:re a0pli¢~l from barr¢t~ :ontaining either sodium t-ttlutama:e (Sigma, 0.5 M, r~H 8.5), S." (i~enirtsula, 2.3 mM in 150raM NaCI) or EJPi)r-sP (Peninsu|a, 3 r a m in tSOmM NaCI, or 3 mM in ?.f)mM sodium acetate, p;-t 4.5). In addition, one barrel of the micropipette contained " M Nat?l for current balancing and current controls, and am-,thtr barrel Pontamine sky blue dye (2,5% in acetate bqffer) for histological verification o~ recording ilr s. DPDT-SP t45 hA, ~ ) was found to dcpre~r, SP respon,~cs (upper trace), t:ut v, hen tested agaiast j °glutarnate.a.voktxt e~citation.,, ( I , lower ~racc), the analogue ',~as found to de!~re',,, tt~esc also.
actic.n,~ when ejected (20--80 nA) onto neurones with a pre-existing ongoing firing .,'ate (eill~er Sl:ontaneous or t-glutamate-evoked), as previously described [13, 15]. When challenged with concurrent applicalion of DPDT-SP (30-150 nA), respons:s to SP were reduced in 6 of 17 neurones (lqg. i). However, in all of these ca.',e,i there v,a:~ also a marked reduction of the ongoing firing rate of th., neurone and/,)r depression of the L-glutamate-evoked activity (Fig. l). Tlr~,e effects of DPDT-SP thus cannot be regarded as examples of specific SP antagonism. On the remaining 11 ncmrones, no depression of SP respon~:s ~ as evident, and the analogue either had no effect (n = 6) or had excitatory actions (n = 5). Excitatory effects tyl~cally consisted of increases in firing rate and paroxysmal 0ursts of firing, which were often ~'ollowed by depression of the neurone. In isolated spinal cord preparations, SP (0. l-2.5 #M) was found to depolarize motoneurones, as previously demribed [9] (Fig. 2). When DPDT-SP was su~rfused at similar concentrations, no pronounced ¢~irect effects were noted, and SP responses in the presence of the analogue were. eithcr similar to or slightly greater than cor, trol responses. When higher concentrations of DPDT-SP were applied (up to 20 #M), direct debolarization of the motoneurones became evident, and SP reslaons,"s :,,ere generally potentiated (Fig. 2). in order to enable a comparison to be. made, between the central and lx'ripheral actions of DPEr'F-SP, the analogue was also tested for SP antagonist action on four
293
frill/i "
SP 7..5,t~W
il
I
r--1 DPDT-SP20~.,t."
~t
[
P-~T-
1
SP
ImV
]
B~M
,.
"
~)mir~
Fig. 2. 0(2 rec~dings o f motoneurone polarization were made from the lumbar ventral roots of .1 hemisected isolated spinal ~ r d preparations taken from 4-g-day-old tats, and drugs we:e appJied by additiort t o the bathing medium [121. The figure shows depolarization~ ~roduced by 2.5 ~M SP superfusion ( I ) before, during and after superfusion with DPDT-SP (~--.21,8 It.M). The analotuc potentiates SP responses, and when applied at 20 uM (D) produces a direct depolarization.
A
--
i DiI~T-SP o
o
i011
o
o
oSP
~----o
'+J+1
//t /
o / / /
/
//(
i,t
/
/
a--~ SP ¢~lone
/
/ +/
0J It
Fig. 3. A: contraction.~ of a guinea pig i!eum to addition of SP (% 2.5 × I0 ~)M ) to the bathin:; mcdiun:. Addition of DPDT-SP (e, I 0 ~' M) al~ o prodttces a contraction, following which re:+ponszs to sir, are con.qd-.rahty redttced. Breaks in the record corr,'s2.~3".d to periods of washing of the ti+,~tte. B: do~--respons¢ curves of I|,e same ileur as in (A) to SP in the r.resence (*) anti abscn:e (31 ¢,f I h " M DPDT-SP. It ,.'an be seen thai the anaiot, ue shifts ~.he SP :esponse curve to the right, and it i,. s,.ll po:,siblc to achieve a roa'~imal SP response in lhe presence or DPD'I-SP.
guinea-pig ileum preparations 11, 31. SP (10 ...`7 M - 5 × :,0 '* M) was found to pro:tuce contractions of the ileum. Addition of DPDT-SP to tt~e batbh~r., medium ( I 0 - ; M - - I O t, M ) was also fc~;nd .to pro~uc,: c.:).;.~r;.,c::..~;n,; :;n+, :,f:e: ~hc.,:c had :mbsided, responses to SP wet'(, reduced (Fig. 3A), as has been described !.~ od~cr ' . swooth muscle preparalions [|0] . The redaction (,f ""~[- ~c"4;,.~:~... Ul;.~,~..~.~~. .'..~b,- ( ;"J / ; to competiliv~ antagonism rathe~ than densensitizati,:~.n, as rna.'.-.:ir:,z:[ ,.:o~+.~r-.cti,.m:,
29~
could still be ~,chieved in the presence of the antagonist by increasing the 5P dose, and the dose-response curve for $P was foead to be shifted to the right by DPDTSP (Fig. 3B), as previously de-.~m"ae~.,[ 1 0 ] , , . . . . , : . . . . SP-induced excitations oi locus ¢o¢~1¢us ~ [ ~ ] . H .o~evet~.em" f i ~ : h ~ < , a . cauda|is and the spinal cord suggest that lids aaaloguedoes ao¢ l m v ¢ ~ ~ properties, or that if it has, that t~cs¢ properties ate masked by other effec'.s of DPDT-SP that appear at concentrations insufficient to produce antagonism. The finding that considerably g:cater amounts of the analogue than of SP are required to produce SP antagonism in smooth mL~cle preparations (Fig. 3) [10| would support this suggest.ion. A further implication of these :¢sults is that recel~OrS for SP in the locus ¢oeruleus may be 4ifferent to those in the spinal cord and n. caudalis, the former being ~:aore sensitive to the antagon:~st actions and less sensitive to the direct effects of DPDT-SP. [t is also noteworthy that DPDT-SP has been fcund to be have neurotoxin-like aclions [7], and it is po~.~sibte that these effects may be related to the direct effects seen in this study. in conclusion, it appears that DPDT-SP is not a suitable tool for the study of putative SP systems in the spinal cord or n. caudalis of the rat, although this cornpound has prov-d u~ful in the peripheral nervous system [10] and in other parts of the central nervous system [2]. Th..~e results provide further evidence for the exi.qt,:ncc of sub-tyl~s o" SP receptor o~ different sites in the body [5, :l]. Wc wi,,h to :hank Dr. R.H. Evan~ for as-,/stanc¢ with the spinal ~:o~:lpreparation. ~t:pr ;rtt:d by gran;:~ froo~ the M.R.C., Royal Society and S.R.C. t [:dml)t+rrb Stati, Phal':~a+<.>]o+Acal l:~;'+;..r;Inel~t.> on i+,ol++t,.xl Pre+'.~+ralion,,. 2l~c+ end., Ci)urch!ll. I.iving,+tc,+,, Edi:+burgh, t9"t+, pp. 2~-2~., 5B-66, 152--153. 2 En,, bcrp.. G., Svc,m,,~'m, T.H., Ro.~¢ll, S. and Felkers, K,+ A ',ynl~+.k: peptich~ as an amaloni+..t of ,~ub~tance P, Nature (I.on~i,L 29~ ~19~.!) 222-223. 3 I'~lk¢,s, K., H6rig, J., Rdsell, 5. and BjO~'ktoth. U.. Chemical desi):n of attlag¢)tli:,l~ t bUbSt&nCf P, ..\eta physic.!. ~cand., ! I ! (19gl, 505-.506. .~ G(,t)¢l, S., Fall~, W.M. and Hockfic~d, S., "l'h¢ divis~.oo of the d o t ~ ! and vei~tfal horn= of lh© mamtn~han caudal medulla inlo eight layers ush~$ anatomk-ai ~:r=;.,,-ria. In D. Andcru~rJ and 13. M.~lthcws (Ed~.), Pain ia =he i'rigemina~ Ro~ion, El~vicr/N,:v:a-H~,~u~d Biomedit~ Pt¢~, An:slc,d=t,~, 1977, ?p. 443-453. '~ }tawco,ck, A.B., Ita],'¢s, A.G. and Ty=.rs, M B.. Aggni,_: prop¢~i¢~, of (t~-P~.o-Phe'r,t~-TrTp *) gLt~',~t;ilZCC P ill vitro - c~i(1¢t~¢¢ t'o~ mote than on¢.ty[~ or substan¢-. P tc~,-~ot, Briz. J. Pharra~.¢.=l., 75 ~t982) l i ~P. 6 Henry. I.I : S~d~.,;:;wce P ~ad poin: an tlod;~ting, "lr~.~ld~, NcuraSci , 3 (l':Yd0) 9~-97. "' t-4(~kfch, | . , Viqc¢~t, S., ~tc~ls,ca, L., Ro~¢ti, S., }'~R,~:~s, K., Marr, cy~ K., Gold,tell1, ~.. ~=tv,J Ct'~c)lc. C.. tmm~i~t~h;.stt×'hemi,,~d ev!denc~ for a "r~*~rozoxk.,' a~;tio~ of (o-Pto2,o-Ttp;.~}-substltrtce P, ~:~ a::a!~,gu¢ ,.~i=h sut'stanc¢ P ~magonistic activity, Ac:a physiol, stand., 113 (198I) 571-573. g P,cr:,cn. L.L.: l.e¢, C.M., ,Land[~:~g, B,E.B. and Watson, S.P., Differins rank orders of ~ t e n c y of ~ubstan¢¢ P a ~alo~lt:¢~ .;u~.(:e,,: ih? ¢.xii!ertce o | multiple reccptct sub.l.vpe.~, P~ti:. J. Ph,tl'~a¢ol.. "/5 (!982) ~.12P.
295 9 Konis~, S. ~ Otzuka, M., Excitatory action of hypothatamic substance P on spinal mote,net, onrs of newborn rats, ~,latur¢ (Lond.). 252 (19"/4) ?34-735.
!0 Leander, $., l-ltlkmson, R., Resell, S., Folkers, K., Sundler, ). and Torp.qvist, K., A .~pccific sub~taace P ~mtalonist l:,k~ckssmooth muscle contractions in.duced by non-choii~.,cL~ic,nonadreneri~ here sfimutatk;n, Nature (Lond.), 294 (I~I) 467-469. 11 Nicoll, R~A,, Sct~.-nk~, C, and L.ceman, S.E., Substance P as a transmitt..'r candidate, Ann. Roy. bleurosci., 3 (1950~ 227-26& 12 Otsuka, M, and Konishi, S., Electrophysiology of mammalian spinal cord in "ilro, Nature (Lond.), 252 (!974) "I33-734. 13 Salt, T.E., Ber,y, 'JlC., Hill, R.G. and Morri:+, R., The effects of p~ptide neuro:ransmitter can~didates on single neuroncs in the rat trigcminal nucleus caudalis. In B. Matthews and R.G. Hill (F:ds.), Anatomical, Fhysiological and Pharmacological Aspects of Tri~cminal Pain, Elsc':icr/NorthHolland Biomcd!cal Press.. Am.~tcrdam, in press. 14 Salt, T.E. and Hill. R.G., Differentiation of excitatory amino acid r.'ccoTor, in h e rat ¢a:~dai trige:ninai nuclc:Js: a microiontophoretic study, Neuroph~rmacology, in prt s',. 15 Zieglgtlnsberger, W. and Tulloch, i.F., Effects of substance P on ncurot,es in the dot-;~~l horP. of th,? spinal cord of t,e cat, Brain Res., 166 (1979) 273-282.