INHIBITION OF NEURONAL HIGH VOLTAGE-ACTIVATED CALCIUM CHANNELS BY INSECT PEPTIDES - A COMPARISON WITH THE ACTIONS OF OMEGA-CONOTOXIN GVIA

The whole cell variant of the patch clamp technique was used to invest igate the actions of two novel insect peptides on high voltage-activat ed Ca2+ currents in cultured dorsal root ganglion (DRG) neurones. The insect peptides (PMP-D2 and PMP-C) were isolated originally from insec t brains and fat bodies, and have been found to have similar three-dim ensional structures to the N-type Ca2+ channel inhibitor omega-conotox in GVIA (omega-CgTx GVIA). High voltage-activated Ca2+ currents were a ctivated from a holding potential of -90 mV by depolarizing step comma nds to 0 mV. Extracellular application of synthetic PMP-D2 or PMP-C (1 mu M) attenuated high voltage-activated Ca2+ currents. The effects of PMP-C were strongly dependent on the frequency of current activation, but inhibition was apparent and reached a steady state after 20 steps when currents were evoked for 30 msec at 0.1 Hz. The actions of the t wo insect peptides overlapped both with each other and with omega-CgTx GVIA, suggesting that N-type Ca2+ current was predominantly sensitive to these peptides. Low voltage-activated T-type current and 1,4-dihyd ropyridine sensitive L-type Ca2+ currents were insensitive to 1 mu M P MP-D2 and PMP-C, which indicates a degree of selectivity. The presence of a fucose group on PMP-C abolished the ability of this peptide to a ttenuate high voltage-activated Ca2+ currents, which may reflect a mec hanism by which peptide function could be regulated in insects. The el ectrophysiological data are supported by studies on Ca-45(2+) influx i nto rat cerebrocortical synaptosomes. Both PMP-D2 (10 mu M), PMP-C (10 mu M) and omega-CgTx GVIA (1 mu M) attenuated a proportion of Ca-45(2 +) influx into the synaptosomes, but additive effects of these peptide s were not observed. We conclude that these naturally occurring peptid es obtained from invertebrate preparations have inhibitory effects on N-type Ca2+ channels. Although the peptides have related three-dimensi onal structures, they have distinct amino acid sequences and appear to have different mechanisms of action to produce inhibition of mammalia n neuronal high voltage-activated Ca2+ channels. (C) 1997 Elsevier Sci ence Ltd.