AMPK inhibits voltage-gated calcium channel-current in rat chromaffin cells.

Metabolic changes are critical in the regulation of Ca2+ influx in central and peripheral neuroendocrine cells. To study the regulation of L-type Ca2+ channels by AMPK we used biochemical reagents and ATP/glucose-concentration manipulations in rat chromaffin cells. AICAR and Compound-C, at low concentration, significantly induce changes in L-type Ca2+ channel-current amplitude and voltage dependence. Remarkably, an overlasting decrease in the channel-current density can be induced by lowering the intracellular level of ATP. Accordingly, Ca2+ channel-current density gradually diminishes by decreasing the extracellular glucose concentration. By using immunofluorescence, a decrease in the expression of CaV1.2 is observed while decreasing extracellular glucose, suggesting that AMPK reduces the number of functional Ca2+ channels into the plasma membrane. Together, these results support for the first time the dependence of metabolic changes in the maintenance of Ca2+ channel-current by AMPK. They reveal a key step in Ca2+ influx in secretory cells.

Modulation of N-type Ca2+ channel current kinetics by PMA in rat sympathetic neurons.

The protein kinase C activator phorbol 12-myristate 13-acetate (PMA) has been used extensively in studies of G protein modulation of Ca2+ channels. PMA has been shown to be a powerful tool for inducing phosphorylation and interrupting G-protein-mediated signaling pathways. Here we re-examine the effects of PMA on whole-cell N-type Ca2+-channel currents in rat sympathetic neurons. We found that, along with an increase in the current amplitude previously reported by others, PMA pretreatment leads to alterations in current activation and inactivation kinetics. These alterations in current kinetics are voltage-dependent and are not reproduced by internal dialysis with the G protein inhibitor GDPbetaS. Alterations in current kinetics by PMA may therefore indicate the existence of a modulated state, presumably phosphorylated, of N-type Ca2+ channels. We propose that the increase in current amplitude is due primarily to alterations in current kinetics rather than to removal of tonic inhibition.

Potentiation of prolactin secretion following lactotrope escape from dopamine action. I. Dopamine withdrawal augments l-type calcium current.

Hypothalamic dopamine (DA) tonically inhibits prolactin (PRL) release from the anterior pituitary gland. Transient escapes from this DA tone elicit a pronounced potentiation of the PRL-releasing action of secretagogues such as thyrotropin-releasing hormone (TRH). Previous evidence has suggested that modulation of Ca(2+) channels can be involved in this potentiation. With a lactotropic cell line (GH(4)C(1)) expressing human D(2)-DA receptors, we tested the hypothesis that a brief escape from the tonic inhibitory action of DA triggers a facilitation of Ca(2+) influx through Ca(2+) channels. We initially found that in these cells, DA effectively and reversibly inhibited PRL secretion, and reversibly enhanced an inwardly rectifying K(+) current. The effects of DA administration and withdrawal on Ca(2+) currents were examined using the patch-clamp technique in the whole-cell configuration and Ba(2+) as a divalent charge carrier through Ca(2+) channels. Macroscopic Ba(2+) currents were significantly decreased by short term (1-10 min) applications of DA (500 nM), which further declined following 24 h of constant exposure to DA. After DA removal, a biphasic facilitation of the density of Ba(2+) currents was observed. An initial 2-fold enhancement of conductance was detected between 10 and 40 min, followed by a second facilitation of the same magnitude observed 24 h after DA withdrawal. The present results directly demonstrate that dissociation of DA from D(2)-receptors expressed in GH(4)C(1) lactotrope cells causes an increase of high-voltage-activated Ca(2+) channel function, which may play an important role in the cross-talking amplification of endocrine cascades such as that involved in the TRH-induced PRL-release potentiating action of DA withdrawal.