Direct effect of Ca super(2+)-calmodulin on cGMP-activated Ca super(2+)-dependent Cl super(-)channels in rat mesenteric artery myocytes

• Published in: The Journal of physiology Vol. 559; no. 2; pp. 449 - 457
• Main Authors: Piper, A S, Large, WA
• Format: Journal Article
• Language: English
• Published: 01.09.2004
• ISSN: 0022-3751

Recently a novel cGMP-activated Ca super(2+)-dependent Cl super(-)channel has been described in rat mesenteric artery smooth muscle cells. In the present work we have investigated the actions of calmodulin (CaM) on single channel cGMP-activated Ca super(2+)-dependent Cl super(-)current (I sub(Cl(cGMP,Ca))) in inside-out patches. When 1 mu M CaM was applied to the intracellular surface of inside-out patches bathed with 10 mu M cGMP and 100 nM [Ca super(2+)] sub(i) there was approximately a 10-fold increase in channel open probability (NP sub(o)). This effect of CaM was not observed with lower [Ca super(2+)] sub(i) and 100 nM [Ca super(2+)] sub(i) with 1 mu M CaM did not activate Cl super(-)channels in the absence of cGMP. The unitary conductance, reversal potential and mean open time of the single-channel currents were similar in the absence or presence of CaM. With 10 mu M cGMP and 100 nM [Ca super(2+)] sub(i) the relationship between NP sub(o) and CaM concentration was well fitted by the Hill equation yielding an equilibrium constant for CaM of about 1.9 nM and a Hill coefficient of 1.7. With 1 mu M CaM (+10 mu M cGMP) the relationship between [Ca super(2+)] sub(i) and NP sub(o) was also fitted by the Hill equation which yielded an apparent equilibrium constant of 74 nM [Ca super(2+)] sub(i) and a Hill coefficient of 4.8. When [Ca super(2+)] sub(i) was increased from 300 nM to 1 mu M there was a decrease in NP sub(o). The potentiating effect of CaM was markedly reduced by the selective CaM binding peptide Trp (5 nM) but not by the Ca super(2+)/CaM-dependent protein kinase II (CaMKII) inhibitor autocamtide II related inhibitory peptide (AIP). It is concluded that CaM potentiates the activity of single channel I sub(Cl(cGMP,Ca)) by increasing the probability of channel opening via a CaMKII-independent mechanism.