REGULATION OF
L-TYPE Ca2 CHANNELS BY PROTEIN KINASE C DURING HYPOXIA IN GUINEA-PIG
VENTRICULAR MYOCYTES. L C Hool.* Physiology
Department, The University of Western Australia, Nedlands, W.A. The L-type Ca2+
channel plays an integral role in cardiac excitation and contraction.
Sympathetic drive is increased during hypoxia. However, little is known about
the effects of b-adrenergic
receptor stimulation on cardiac L-type Ca2+ conductance (ICa)
during hypoxia. The effects of hypoxia on ICa in the absence and
presence of the b-adrenergic
receptor agonist isoproterenol (Iso) were examined using the whole-cell
patch-clamp technique. ICa was determined as the peak inward current
recorded during the voltage step to O mV. Exposing myocytes to hypoxia resulted
in a reversible inhibition (25.2 ±
2.5%, n = 28) of basal ICa. In addition hypoxia significantly
decreased the Ko.5 for activation of ICa by Iso from 5.3 ± 0.7 to 1.6 ± 0.08 nM. To determine if the effects were
membrane-delimited, cells were exposed to the membrane -impermeant
thiol-specific oxidising compound DTNB. In the presence of 200 mM DTNB, the inhibition of basal ICa
by hypoxia was attenuated 81.3 ± 9.4%
(n=5) while the increase in sensitivity of 1Ca to Iso was unchanged.
The thiol-specific reducing agent DTT (1 mM) mimicked the effects of hypoxia on
basal ICa and
the response of ICa to Iso. This suggests that the mechanism for the
effects involved the reduction of thiol groups on an effector protein and /or
the channel itself. It has been shown that hypoxia can increase the
translocation of protein kinase C (PKC) - b isoform in the heart. A possible role for the
second messenger in the increase in sensitivity of ICa to Iso during
hypoxia was investigated. Cells were exposed to the PKC inhibitor
bisindolymaleimide 1 HCI (300 nM) during hypoxia and ICa measured in response to increasing
concentrations of Iso. The PKC inhibitor significantly attenuated the increase
in sensitivity of ICa to Iso. Similar results were obtained when
cells were dialyzed with the PKC-b
isoform peptide that prevents the translocation and function of the Ilozyme.
These data provide novel evidence for the regulation of L-type Ca2+
channels by PKC during hypoxia.
