Modulation of Ca(2+)-channel currents by protein kinase C in adult rat sympathetic neurons

Abstract

1. Modulation of Ca(2+)-channel currents by phorbol-12-myristate-13-acetate (PMA) was investigated in acutely dissociated adult rat superior cervical ganglion neurons using the whole cell variant of the patch-clamp technique. 2. PMA (500 nM) increased the current amplitudes, accelerated the inactivation of step currents, retarded the deactivation of tail currents, and shifted the tail current activation to more negative potentials. 3. The effects of PMA were concentration and voltage dependent and mediated through activation of protein kinase C (PKC). PMA also increased Ca2+ currents recorded with the perforated patch technique. 4. PMA affected the N-type Ca2+ channels and an omega-conotoxin GVIA-resistant current component. Ca2+ currents affected by PMA were not sensitive to omega-agatoxin IVA or nimodipine. 5. PMA not only attenuated Ca(2+)-channel inhibition induced by alpha 2-adrenoceptor agonist, which modulates Ca2+ channels via a pertussis toxin (PTX)-sensitive pathway, but also attenuated current inhibition by vasoactive intestinal polypeptide, which modulates Ca2+ channels via a PTX-insensitive but cholera toxin-sensitive pathway. 6. PMA reversed Ca(2+)-channel inhibition induced by tonic activation of G-protein in the absence of neurotransmitter (even in neurons pretreated with PTX) or induced by activation of G-proteins with guanosine 5'-O-(3-thiotriphosphate) (GTP)-gamma-S. 7. Inhibition of phosphatase by okadaic acid or substitution of Ba2+ for Ca2+ in the external solutions accelerated the PMA effect. 8. Our results suggest that activation of PKC antagonizes G-protein mediated inhibition of Ca2+ channels by shifting Ca2+ channels from the "reluctant" state to the "willing" state. The G-proteins and, more likely, the N-type Ca2+ channels may be the target of PKC phosphorylation. Protein phosphatases may be involved in counteracting the PKC phosphorylation in rat sympathetic neurons.

Exogenous nerve growth factor attenuates opioid-induced inhibition of voltage-activated Ba2+ currents in rat sensory neurons

“…In some experiments, reversal of G protein-mediated inhibition was examined using a voltage protocol featuring two steps of 20 ms duration to a test potential of −10 mV, separated by a depolarizing pulse of 40 ms duration to a potential of +70 mV. Voltage protocols similar to this have previously been used to demonstrate relief of G protein-mediated inhibition (Swartz, 1993;Zhu and Ikeda, 1994;Herlitze et al, 1996;Zhu and Yakel, 1997). …”

Up‐regulation of L‐ and non‐L, non‐N‐type Ca2+ channels by basal and stimulated protein kinase C activation in insulin‐secreting RINm5F cells

“…In agreement with this, stimulation of PKC by acute application of phorbol ester causes only minor up-regulation of both channel subtypes, with a net prevalence of the L-type. Since PKC-induced phosphorylation has been proved to disrupt neurotransmitter inhibition on Ca 2+ channels in different neurons and secretory cells [13][14][15][16], we also tested its direct involvement in the Gprotein-mediated noradrenergic inhibition of Ca 2+ channels [17]. The present findings exclude any relation between noradrenergic and PKC-mediated Ca 2+ channels modulation, but underline the essential role of PKC-dependent phosphorylation in tonically maintaining voltage-dependent Ca 2+ channels in a functional state.…”

“…Support to this hypothesis comes from studies in central and peripheral neurons [12,13,15,23], where PKC activation is shown to increase Ca 2+ currents by removing the Ca 2+ channels inhibition induced by activated G proteins either tonically or through neurotransmitter receptors. We found, however, that this is not the case.…”

“…We also provide evidence that PKC-mediated phosphorylation is not involved in the G-protein-mediated noradrenergic modulation of Ca 2+ channels in RINm5F cells. channels are also implicated in insulin release from human 13 cells [10] and catecholamines secretion from chromaffin cells [11] and that they can be up-regulated by PKC activation in neurons [12,13], PKC-dependent phosphorylation of these channels in RINm5F cells may be relevant to the overall secretory activity of these cells.…”

“…We also provide evidence that PKC-mediated phosphorylation is not involved in the G-protein-mediated noradrenergic modulation of Ca 2+ channels in RINm5F cells. channels are also implicated in insulin release from human 13 cells [10] and catecholamines secretion from chromaffin cells [11] and that they can be up-regulated by PKC activation in neurons [12,13], PKC-dependent phosphorylation of these channels in RINm5F cells may be relevant to the overall secretory activity of these cells.Here we show that both L-and non-L, non-N-type Ca 2+ channels are effectively up-regulated by PKC-dependent phosphorylation under basal conditions. In agreement with this, stimulation of PKC by acute application of phorbol ester causes only minor up-regulation of both channel subtypes, with a net prevalence of the L-type.…”