Abstract
Cardiac myocyte contractility is initiated by Ca2+ entry through the voltage-dependent L-type Ca2+ channel (LTCC). To study the effect of Galpha q on the cardiac LTCC, we utilized two transgenic mouse lines that selectively express inducible Galpha q-estrogen receptor hormone-binding domain fusion proteins (Galpha qQ209L-hbER or Galpha qQ209L-AA-hbER) in cardiac myocytes. Both of these proteins inhibit phosphatidylinositol (PI) 3-kinase (PI3K) signaling, but Galpha qQ209L-AA-hbER cannot activate the canonical Galpha q effector phospholipase Cbeta (PLCbeta). L-type Ca2+ current (I(Ca,L)) density measured by whole-cell patch clamping was reduced by more than 50% in myocytes from both Galpha q animals as compared with wild-type cells, suggesting that inhibition of the LTCC by Galpha q does not require PLCbeta. To investigate the role of PI3K in this inhibitory effect, I(Ca,L) was measured in the presence of various phosphoinositides infused through the patch pipette. Infusion of PI 3,4,5-trisphosphate (PI(3,4,5)P3) into wild-type myocytes did not affect I(Ca,L), but it fully restored I(Ca,L) density in both Galpha q transgenic myocytes to wild-type levels. By contrast, PI 4,5-bisphosphate (PI(4,5)P2) or PI 3,5-bisphosphate had no effect. Infusion with p110beta/p85alpha or p110gamma PI3K in the presence of PI(4,5)P2 also restored I(Ca,L) density to wild-type levels. Last, infusion of either PTEN, a PI(3,4,5)P3 phosphatase, or the pleckstrin homology domain of Grp1, which sequesters PI(3,4,5)P3, reduced the peak I(Ca,L) density in wild-type myocytes by approximately 30%. Taken together, these results strongly suggest that the inhibitory effect of Galpha q on the cardiac LTCC is mediated by inhibition of PI3K.
Highlights
Cardiac myocyte contractility is initiated by Ca2؉ entry through the voltage-dependent L-type Ca2؉ channel (LTCC)
Using transgenic mice that selectively express an inducible G␣qQ209L protein in cardiac myocytes, we have demonstrated that activation of G␣q leads to inhibition of the cardiac ICa,L.4
Effects of PI[3,4,5]P3 on ICa,L in Cardiac Myocytes—In this study we employed myocytes isolated from two transgenic mouse lines that selectively express silent G␣q proteins in the heart
Summary
Cardiac myocyte contractility is initiated by Ca2؉ entry through the voltage-dependent L-type Ca2؉ channel (LTCC). Activation of these G␣q proteins in response to injection with tamoxifen, which is converted to 4-HT in animals, causes a large reduction in ICa,L density in cardiac myocytes from both transgenic animals.4 This result suggests that inhibition of the cardiac LTCC by G␣q occurs independently of PLC and may be due to reduced PI3K signaling.
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