Abstract 13760: A Caveolin Targeted L-type Ca2+ Channel Antagonist Inhibits Hypertrophic Signaling Without Reducing Contractility of Cardiac Myocytes
The Ca2+ that activates pathological hypertrophy is not thought to be the same Ca2+ that activates contraction. We hypothesize that Ca2+ influx through a subpopulation of L-type Ca2+ channels (CaV1.2; ICa,L) localized in caveolin (Cav) containing membrane signaling microdomains locally activates Ca2+/calmodulin and calcineurin-mediated NFAT nuclear translocation to induce pathological hypertrophy. To construct a Cav targeted CaV1.2 blocker we modified Rem-GTPase, a protein that potently inhibits CaV1.2. We truncated the membrane association c-terminus of Rem (Rem1-265) which is required for CaV1.2 inhibition and fused Rem1-265 to a canonical caveolin binding domain to create Rem1-265-Cav.
Results: Adenoviral-mediated expression of wild-type Rem (WT-Rem) in adult feline ventricular myocytes inhibited >90% of ICa,L while Rem1-265 had no effects. Rem1-265-Cav caused a small inhibition (less than 15%) of ICa,L . Myocytes infected with WT-Rem had markedly reduced fractional shortening (1.1+/-0.1% resting cell length) while those infected with truncated Rem1-265 (5.0+/-0.6%) and Rem1-265-Cav (4.4+/-0.5%) had contractions not significantly smaller than uninfected myocytes (4.7+/-0.5%). Myocytes expressing Rem1-265-Cav responded normally to isoproterenol while those infected with WT-Rem failed to respond. The β2-specific agonist Zinterol increased ICa,L in control but not in Rem1-265-Cav infected myocytes, consistent with the idea that β2-adrenergic signaling modulates the activity of CaV1.2 within caveolae. Sucrose density gradient fractionation revealed that Rem1-265-Cav cosedimented exclusively with Cav-3 enriched low-density fractions and did not disrupt normal CaV1.2 caveolar targeting. Cells were paced for 1 hour at 1 Hz to induce NFAT-GFP nuclear translocation (hypertrophic signaling: measured as the nuclear to cytoplasmic NFAT-GFP ratio). WT-Rem fully inhibited pacing induced nuclear NFAT translocation and truncated Rem1-265 had no effect. Rem1-265-Cav inhibited more than 90% of pacing induced NFAT nuclear translocation.
Conclusion: CaV1.2 in Cav-3 signaling microdomains are a major source of hypertrophic Ca2+ signaling and can be selectively blocked by Rem1-265-Cav without altering cardiac contractility.
- © 2011 by American Heart Association, Inc.