Calcium-induced calcium release in neurosecretory insect neurons:fast and slow responses

Abstract

The dynamics of intracellular free Ca2+([Ca2+]i) changes were investigated in dorsal unpaired median (DUM) neurons of the cockroach Periplaneta americana. Activation of voltage-gated Ca2+channels caused a steep increase in [Ca2+]i. Depolarizations lasting for < 100ms led to Ca2+release from intracellular stores as is indicated by the finding that the rise of [Ca2+]iwas greatly reduced by the antagonists of ryanodine receptors, ryanodine and ruthenium red. There is a resting Ca2+current which is potentiated on application of a neuropeptide, Neurohormone D (NHD), a member of the adipokinetic hormone family. Ca2+influx enhanced in this way again caused a rise of [Ca2+]isensitive to ryanodine and ruthenium red. Such rises developed and relaxed much more slowly than the depolarization-induced signals. Ca2+responses similar to those induced by NHD were obtained with the ryanodine receptor agonists caffeine (20mM) and cADP-ribose (cADPR, 100nM). These Ca2+responses, however, varied considerably in size and kinetics, and part of the cells did not respond at all to caffeine or cADPR. Such cells, however, produced Ca2+rises after having been treated with NHD. Thus, the variability of Ca2+signals might be caused by different filling states of Ca2+stores, and the resting Ca2+current seems to represent a source to fill empty Ca2+stores. In line with this notion, block of the endoplasmic Ca2+pump by thapsigargin (1μM) produced either no or largely varying Ca2+responses. The Ca2+signals induced by caffeine and cADPR displayed different sensitivity to ryanodine receptor blockers. cADPR failed to elicit any response when ryanodine or ruthenium red were present. By contrast, the response to caffeine, in the presence of ryanodine, was only reduced by about 50% and, in the presence of ruthenium red, it was not at all reduced. Thus, there may be different types of Ca2+release channels. Block of mitochondrial Ca2+uptake with carbonyl cyanidem -chlorophenylhydrazone (CCCP, 1μM) completely abolished cADPR-induced Ca2+signals, but it did not affect the caffeine-induced signals. Taken together our findings seem to indicate that there are different stores using different Ca2+uptake pathways and that some of these pathways involve mitochondria.