Study for the effects of neuroactive peptides newly isolated

本研究采用电生理学方法，对生物活性肽对非洲大蜗牛（Achatina fulica Ferussac）可识别的巨神经元的影响进行了全面的研究。而对哺乳类神经递质肽类如P物质、甲硫氨酸脑啡肽、神经降压素等不敏感，除了催产素另一方面，这些神经元对从无脊椎动物中分离的神经活性肽敏感。这些结果与小分子神经递质（如多巴胺、5-羟色胺和GABA）的结果不同，认为一种肽类神经递质对动物神经元的作用程度比小分子神经递质小。我们与三得利生物有机研究所和圣托马斯大学合作从Achatina神经节中分离出achatin-I（Gly-D-Phe-Ala-Asp），并对Achatin-I的兴奋作用进行了研究。APGW-酰胺（Ala-Pr 关于我们 o-Gly-Trp-NH_2）也是我们从Achatina神经节中分离到的被抑制的Achatina巨神经元。这两种肽被认为是Achatina神经元的神经递质。此外，我们还发现另一组从Achatina神经节中分离的Achatina cardiac-excitatory peptide-1（Ser-Gly-Gln-Ser-Trp-Arg-Pro-Gln-Gly-Arg-Phe-NH_2）对Achatina巨神经元有兴奋作用，而另一组从Achatina神经节中分离的Fulicin（Phe-D-Asn-Gln-Phe-Val-NH_2）对这些神经元有抑制作用。我们还证明了FMRFFamide（Phe-Met-Arg-Phe-NH_2），AF1从蛔虫中分离的FMRFamide的相关化合物（Lys-Asn-Glu-Phe-Ile-Arg-phe-NH_2）和[Ser^2]-贻贝抑制肽（[Ser^2]-MIP）（Gly-Ser-Pro-Met-Phe-Val-NH_2），对achatin-I兴奋作用的细胞内信号转导系统和fulicin抑制作用的细胞内信号转导系统进行了电生理学研究。H-89（蛋白激酶（PK）A抑制剂）和W-7（钙调素抑制剂）可抑制achatin-I的作用，IBMX（环AMP磷酸二酯酶抑制剂）可促进achatin-I的作用，表明achatin-I通过环AMP-PKA系统发挥作用，钙调素参与了该系统。而KT-5823（PKG抑制剂）、calphostin C（PKC抑制剂）和W-7则可抑制Fulicin的抑制作用，表明Fulicin通过环GMP-PKG系统和/或IP_3和DG-PKC系统发挥作用，钙调素参与了这些系统。IBMX可抑制这种抑制作用，其机制目前尚不清楚。我们证明APGW-酰胺和achatin-I对Achatina神经元不仅是一种神经递质，而且也是一种神经调质。少

This year, we studied comprehensively effects of biologically-active peptides on the identifiable giant neurones of an African giant snail (Achatina fulica Ferussac) by the electrophysiological methods. The Achatina giant neurones were insensitive to the mammalian neurotransmitter peptides, such as substance P,Met-enkephalin, neurotensin, etc., except for oxytocin. On the other hand, these neurones were sensitive to the neuroactive peptides isolated from invertebrates. These findings were different from those of small molecule putative neurotransmitters, such as dopamine, serotonin and GABA.With these results, it was considered that one peptide neurotransmitter acts on the neurones of the less extent of animal species than the small molecule putative neurotransmitter. Achatina giant neurones were excited by achatin-I (Gly-D-Phe-Ala-Asp), isolated from Achatina ganglia by the collaboration of us, Suntory Institute for Bioorganic Research and University of Santo Tomas. APGW-amide (Ala-Pr … More o-Gly-Trp-NH_2), isolated also from Achatina ganglia by us, were inhibited Achatina giant neurones. These two peptides were proposed to be neurotransmitters for Achatina neurones. Besides, we demonstrated that Achatina cardio-excitatory peptide-1 (ACEP-1) (Ser-Gly-Gln-Ser-Trp-Arg-Pro-Gln-Gly-Arg-Phe-NH_2) isolated from Achatina ganglia by another group showed excitatory effects on Achatina giant neurones, and that fulicin (Phe-D-Asn-Gln-Phe-Val-NH_2) isolated from Achatina ganglia by another group had inhibitory effects on these neurones. We demonstrated also that FMRFFamide (Phe-Met-Arg-Phe-NH_2) isolated from the clam ganglia, AF1 (Lys-Asn-Glu-Phe-Ile-Arg-phe-NH_2), a related compound of FMRFamide isolated from an Ascaris, and [Ser^2]-Mytilus inhibitory peptide ([Ser^2]-MIP) (Gly-Ser-Pro-Met-Phe-Val-NH_2) isolated from Mytilus, had inhibitory effects on these neurones.The intracellular signal transduction systems for the excitatory effects of achatin-I and the inhibitory effects of fulicin were studied electrophysiologically using the inhibitors for these systems. H-89 (protein kinase (PK) A inhibitor) and W-7 (calmodulin inhibitor) inhibited the achatin-I effects, and IBMX (cyclic AMP phosphodiesterase inhibitor) facilitated the same effects, indicating that achatin-I acted via cyclic AMP-PKA system, and calmodulin participated in this system. On the other hand, KT-5823 (PKG inhibitor), calphostin C (PKC inhibitor) and W-7 suppressed the inhibitory effects of fulicin, indicating that fulicin acted via cyclic GMP-PKG system and/or IP_3 and DG-PKC systems, and calmodulin participated in these systems. IBMX suppressed the inhibitory effects, the mechanism of which cannot be explained at present.We demonstrated that APGW-amide, as well as achatin-I,acted not only as a neurotransmitter but also as a neuromodulator for Achatina neurones. Less