CHARACTERIZATION OF SUBSTANCE-P RELEASE FROM THE INTERMEDIATE AREA OFRAT THORACIC SPINAL-CORD

Substance P (SP) nerve terminals innervate the intermediolateral cell column (IML) of the thoracic spinal cord, where SP coexists with serot onin (5-HT), neurokinin A (NKA) and thyrotropin-releasing hormone (TRH ). Neither the depolarization-induced release of SP nor the presence o f other neurochemicals in the regulation of SP release has been direct ly studied in this system. In the present study, basal and K+-stimulat ed release of SP from the microdissected intermediate area (including the IML, intercalated nucleus and central autonomic nucleus) of the ra t thoracic spinal cord, and the regulation of SP release by presynapti c autoreceptors and by coexisting neurochemicals (5-HT, NKA and TRH) w ere studied using an in vitro superfusion system. Potassium evoked a c oncentration- and extracellular Ca2+-dependent release of SP. In rats pretreated with the serotoninergic neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT), both SP content and the absolute amount of SP released wer e decreased. However, the fraction of the remaining tissue content of SP released by K+ depolarization was not changed subsequent to 5,7-DHT treatment. Moreover, 5-HT, 5-HT1B agonists (CGS-12066B and RU 24969) and a 5-HT3 agonist (2-methyl-5-HT) did not alter the K+-evoked releas e of SP. These data demonstrate that SP is released from the intermedi ate area of the rat thoracic spinal cord and some of the SP released c omes from serotoninergic nerve terminals. Although 5-HT coexists with SP in the IML, neither endogenous 5-HT nor 5-HT receptor ligands appea r to regulate the release of SP. Other colocalized neuropeptides (NKA and TRH) are not involved in the regulation of SP release because neit her NKA, a NK2 agonist (GR 64349) nor a TRH analog (MK-771) changed th e K+-evoked release of SP. A neurokinin-1 (NK1) antagonist (GR 82334) dose-dependently (10(-9)-10(-7) M) increased the K+-stimulated release of SP. These data suggest the presence of presynaptic inhibitory NK1 autoreceptors. Whereas, NK(?)1 agonists, [GR 73632 (10(-9)-10(-6) M) a nd [Sar(9), Met (O-2)(11)]SP (10(-8)-10(-6) M)], increased the basal a nd K+-stimulated release of SP, the excitatory effects of GR 73632 wer e not blocked by the NK1 antagonist. Moreover, GR 73632 increased the efflus of SP to a greater extent in the absence of peptidase inhibitor s. Thus, the effect of NK1 agonists on the release of SP may be relate d to an inhibition of peptide degradation rather than activation of NK 1 autoreceptors. (C) 1996 Wiley-Liss, Inc.