Evidence for a Role for the Cytoskeleton in Communication Between the L-Type Calcium Channel and the Mitochondria in Isolated Cardiac Myocytes

Abstract

Cytoskeletal proteins stabilize cell structure but also regulate subcellular distribution of mitochondria and cardiac L-type Ca2+ channel (LTCC) activity. We have previously demonstrated that mitochondrial function can be regulated by alterations in LTCC activity. This effect was attenuated when the cytoskeleton was disrupted with latrunculin A. To further explore this, we determined whether regulation of mitochondrial function by the LTCC is altered in a murine model of Duchenne Muscular Dystrophy(mdx). Mitochondrial membrane potential (Ψm) and metabolic activity was assessed after activation of the LTCC in cardiac myocytes isolated from C57BL/10ScSn-Dmdmdx/Arc (mdx) and C57BL/10ScSnArc (control) mice. Exposure of control myocytes to 10μM BayK(-) (LTCC agonist) caused a 11.4±1.7% increase in Ψm assessed as alterations in JC-1 compared to myocytes exposed to inactive BayK(+) (n=8, p<0.05). The response was attenuated when myocytes were exposed to LTCC antagonist nisoldipine (n=7). However BayK(-) did not induce any significant alteration in JC-1 signal in myocytes from mdx mice (n=6). In control myocytes BayK(-) caused a 105.4±7.4% increase in metabolic activity assessed using an MTT assay (n=8, p<0.05). The response was attenuated when myocytes were exposed to nisoldipine (n=8) or mitochondrial calcium uniporter inhibitor Ru360 (n=8) but unaltered when exposed to ryanodine receptor antagonist dantrolene (n=4). Exposure of mdx myocytes to BayK(-) did not induce any significant alteration in metabolic activity (n=8). These data confirm that alterations in LTCC activity can modulate mitochondrial function and that the cytoskeleton plays an important role in mediating this response. Since the LTCC is the initiator of contraction it has been proposed that a functional coupling between the LTCC and mitochondria may assist in meeting myocardial energy demand on a beat to beat basis.