ABSTRACT
Intracellular recordings were made from identified neurons from the suboesophageal ganglia of Helix aspersa. The inhibitory action of nine S-Iamide peptides was investigated. Structure-activity studies suggest that all act through a common receptor, which normally requires FVRIamide at the C terminal, with a preferred length of seven amino acids. Substitution at the N-terminal with alanine (A), threonine (T), proline (P) or leucine (L) results in little change in potency, suggesting the N-terminal requirements are relatively flexible. Ion substitution experiments suggest that potassium is the main ion involved in the inhibitory response to S-Iamide application. Studies using a range of compounds, which modify second messenger systems, would suggest that S-Iamide peptides may interact with adenylate cyclase. No evidence was found for an interaction with either guanylate cyclase or nitric oxide synthase.
Subject(s)
Adenine/analogs & derivatives , Helix, Snails/physiology , Neurons/drug effects , Neuropeptides/chemistry , Neuropeptides/pharmacology , 1-Methyl-3-isobutylxanthine/pharmacology , Adenine/pharmacology , Adenylyl Cyclase Inhibitors , Amides/chemistry , Amides/pharmacology , Amino Acid Sequence , Animals , Chlorides/pharmacology , Colforsin/pharmacology , Cyclic AMP/biosynthesis , Dose-Response Relationship, Drug , Electrophysiology , Enzyme Inhibitors/pharmacology , Neurons/physiology , Phosphodiesterase Inhibitors/pharmacology , Potassium/pharmacology , Second Messenger Systems , Structure-Activity RelationshipABSTRACT
Intracellular recordings were made from identified neurones in the suboesophageal ganglia of the snail, Helix aspersa. The actions of the eight FxRFamide analogues were investigated on these neurones. These peptides included ones isolated from arthropods and nematodes. All the peptides excited certain neurones while inhibiting others, though their relative potencies varied. Overall on neurones inhibited by these peptides the potency order was: DNFLRFamide > FMRFamide > PDVDHVFLRFamide = KNEFIRFamide > FLRFamide >> SDRNFLRFamide = SDPNFLRFamide > KHEYLRFamide. However, if the responses are compared on individual cell types, then the picture becomes more complex. For example, on cell F-2, KNEFIRFamide proved to be potent with an EC-50 value of 0.54 microM. On neurones F-13/16 and E-16, PDVDHVFLRFamide was inhibitory while FMRFamide, FLRFamide, SDRNFLRFamide and SDPNFLRFamide were excitatory. In terms of overall excitatory actions, the data are less complete but an approximate order of potency is: FMRFamide > DNFLRFamide >> SDPNFLRFamide > PDVDHVFLRFamide >> KNEFIRFamide = KHEYLRFamide = SDRNFLRFamide. However this again varies between specific neurones. These results demonstrate that peptides from insects, crustacea and nematodes are active on Helix neurones and may activate specific receptor subtypes, indicating the possible presence of endogenous analogues of these non-molluscan peptides in the Helix nervous system.
Subject(s)
Neurons/drug effects , Neuropeptides/pharmacology , Amides/chemistry , Amino Acid Sequence , Animals , Dose-Response Relationship, Drug , Helix, SnailsABSTRACT
Intracellular recordings were made from neurones E-8, E-16 and E-13a in the visceral ganglion of Helix aspersa. GSPYFVamide inhibits the activity of these neurones and the role of a second messenger system in this inhibition was investigated. 8-Bromo-cGMP, 100 microM was found to potentiate this inhibition while ODQ, 100 microM, an inhibitor of guanylyl cyclase, almost completely blocked GSPYFVamide-induced inhibition. Four NO donors sodium nitroprusside, 100 microM, sodium nitrite, 1 mM, SNOG, 50 microM, and SNAP, 10-50 microM, all potentiated the GSPYFVamide-induced inhibition. L-NAME, 100-1000 microM, a competitive inhibitor of NOS, blocked the GSPYFVamide-induced inhibition. In some cases recovery was only partial. The possible role of NO in modulating the inhibitory response to GSPYFVamide is discussed.
Subject(s)
Neurons/metabolism , Nitric Oxide/metabolism , Oligopeptides/pharmacology , Animals , Cyclic GMP/analogs & derivatives , Cyclic GMP/pharmacology , Cyclic GMP-Dependent Protein Kinases/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Glutathione/analogs & derivatives , Glutathione/pharmacology , Guanylate Cyclase/antagonists & inhibitors , Helix, Snails , NG-Nitroarginine Methyl Ester/pharmacology , Neurons/drug effects , Neurons/physiology , Nitroprusside/pharmacology , Nitroso Compounds/pharmacology , Oligopeptides/metabolism , Oxadiazoles/pharmacology , Penicillamine/analogs & derivatives , Penicillamine/pharmacology , Quinoxalines/pharmacology , S-Nitrosoglutathione , Sodium Nitrite/pharmacologyABSTRACT
1. The structural and ionic requirements for potent FVamide-induced inhibition were investigated using Helix aspersa central neurons under current or voltage clamp. 2. For potent FVamide inhibition the full hexapeptide sequence, GSPYFVamide, was required. The rank order of decreasing potency was GSPYFVamide >> SPYFVamide> PYFVamide > YFVamide. 3. GSPYFVamide inhibition involved an increase in conductance to both potassium and chloride ions as demonstrated by ion substitution experiments and addition of 4-aminopyridine, tetraethylammonium, 4,4-di-isothiocyanatostilbene-2,2'-disulfonic acid, 4,4-dinitrostilbene-2,2'-disulfonic acid disodium salt and furosemide to the solution bathing the preparation.
