Ghrelin-induced growth hormone release from goldfish pituitary cells involves voltage-sensitive calcium channels.
Ghrelin (GRL) is a stimulator of growth hormone (GH) release in many organisms, including goldfish. As a first study to examine the signalling mechanisms mediating GRL action on GH release in goldfish, we tested the hypothesis that GLR induces GH release from goldfish pituitary cells by enhancing Ca(2+) entry through L-type voltage-sensitive Ca(2+) channels (LVSCCs) using perifusion GH release and fura-2/AM Ca(2+)-imaging experiments. Goldfish (g)GRL(19) at 1 nM elicited reversible and repeatable GH responses from dispersed goldfish mixed pituitary cultures. However, the lack of a dose-response relationship in sequential treatments with decreasing concentrations of gGRL(19) (ranging from 10 to 0.01 nM) implicated rapid desensitization of the GH response. Sequential applications of gGRL(19) (1 nM) and salmon GnRH (100 nM), a known Ca(2+)-dependent stimulator of GH release, increased intracellular free Ca(2+) levels ([Ca(2+)](i)) from the same identified somatotropes, suggesting co-expression of GRL and GnRH receptors on single cells. In contrast, 1 nM gGRL(19) failed to elicit GH release and elevation in [Ca(2+)](i) when the cells are incubated with nominally Ca(2+)-free media. When GH release and [Ca(2+)](i) increases were already stimulated by the LVSCC agonist Bay K8644 (10 microM), addition of 1 nM gGRL(19) did not further elevate these responses. Finally, the LVSCC inhibitors nifedipine (1 microM) and verapamil (1 microM) abolished 1nM gGRL(19)-induced GH release responses while nifedipine eliminated gGRL(19)-induced [Ca(2+)](i) increase. Taken together, the results of this study provide evidence that entry of extracellular Ca(2+) through LVSCCs is a key component of the GRL signalling pathway leading to GH release in the goldfish pituitary.
Grey
Caleb L
CL
Department of Biological Sciences, University of Alberta, Edmonton, Alta., Canada T6G 2E9.
Chang
John P
JP
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002120:Calcium Channel Agonists / Q000494:pharmacology; D015220:Calcium Channels / Q000378:metabolism*; D002478:Cells, Cultured / Q000378:metabolism*; D054439:Ghrelin / Q000494:pharmacology*; D006054:Goldfish; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology; D013006:Growth Hormone / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D010902:Pituitary Gland / Q000166:cytology*
D002120:Calcium Channel Agonists; D015220:Calcium Channels; D054439:Ghrelin; D007987:Gonadotropin-Releasing Hormone; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D013006:Growth Hormone; D009543:Nifedipine; D002118:Calcium