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Department of Physiology and Biophysics, University of Nebraska Medical Center, Omaha, Nebraska 68198-4575
Activation of mesangial cells by ANG II provokes release of intracellular Ca2+ stores and subsequent Ca2+ influx through voltage-gated channels, events that are reflected by a large transient increase in intracellular concentration [Ca2+]i followed by a modest sustained elevation in [Ca2+]i. These ANG II-induced alterations in [Ca2+]i elicit activation of large Ca2+-activated K+ channels (BKCa) in a negative-feedback manner. The mechanism of this BKCa feedback response may involve the direct effect of intracellular Ca2+ on the channel and/or channel activation by regulatory enzymes. The present study utilized patch-clamp and fura 2 fluorescence techniques to assess the involvement of multifunctional calcium calmodulin kinase II (CAMKII) in the BKCa feedback response. In cell-attached patches, KN62 (specific inhibitor of CAMKII) either abolished or reduced to near zero the ANG II-induced BKCa feedback response. This phenomenon did not reflect direct effects of KN62 on the BKCa channel, because this agent alone did not significantly alter BKCa channel activity in inside-out patches. KN62 also failed to alter either the transient peak or sustained plateau phases of the [Ca2+]i response to ANG II. In inside-out patches (1 µM Ca2+ in bath), calmodulin plus ATP activated BKCa channels in the presence but not the absence of CAMKII. These observations are consistent with the postulate that CAMKII is involved in the BKCa feedback response of mesangial cells, acting to potentiate the influence of increased [Ca2+]i on the BKCa channel or a closely associated regulator of the channel. An additional effect of CAMKII to activate a voltage-gated Ca2+ channel cannot be ruled out by these experiments.
mesangial cells; angiotensin II; patch clamp; fura 2
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