Effects of Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) on Intracellular cAMP and Ca²⁺ Levels in Common Carp (Cyprinus carpio) Pituitary Cells in vitro

Abstract

In
order to study signal transduction in PACAP-stimulated GH releas e , effects
of ovine PACAPs (oPACAPs) on cAMP synthesis and intracellular free Ca²⁺([Ca²⁺]i)
levels in primarily cultured common carp (Cyprinus carpio) pituitary
cells were investigated using radioimmunoassay (RIA) for cAMP and [Ca²⁺]i
Indo-1/AM loading techniques. The main results obtained could be briefly summarized
as follows: parallel with their GH-releasing effects, oPACAP-38 (0.01 nmol/L
to 1 μmol/L) and oPACAP-27 (0.01 nmol/L to 1 μmol/L) were also effective in
increasing cAMP release, cellular cAMP content, and total cAMP production
in a dose-dependent manner in common carp pituitary cells. In addition to
their cAMP synthesis , 0.5 μmol/L oPACAP-38 and 0.5 μmol/L oPACAP-27 were
also effective in elevating [Ca²⁺]i levels in common
carp pituitary cells preloaded with the Ca²⁺-sensitive dye Indo-1;
the stimulatory actions were dependent on the presence of extracel lular Ca²⁺
([Ca²⁺]e), as removal of [Ca²⁺]e
by adding the Ca²⁺ chelator 4 mmol/L E GTA blocked the rise of
[Ca²⁺]i induced by oPACAP-38 and oPACAP-27, respectively.
Besides, the rise in [Ca²⁺]i induced by 0.5 μmol/L oPACAP-38
was blocked by treatment with 10 μmol/L nifedipine, an inhibitor of “L-type”
voltage-sensitive Ca²⁺ channels (VSCC); but 0.5 μmol/L oPACAP-38
was not effective in elevating [Ca²⁺]i levels after
the cells treated with Ca²⁺-sensitive dye Indo-1 were challeng
ed with 10 μmol/L nifedipine. These results, taken as a whole, strongly suggest
the involvement of the cAMP- and Ca²⁺-dependent pathways in PACAP-stimulated
GH release in common carp.