Abstract
We recently demonstrated that a 17-ketosteroid, epiandrosterone, attenuates L-type Ca2+ currents (ICa-L) in cardiac myocytes and inhibits myocardial contractility. Because 17-ketosteroids are known to inhibit glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme in the pentose phosphate pathway, and to reduce intracellular NADPH levels, we hypothesized that inhibition of G6PD could be a novel signaling mechanism which inhibit ICa-L and, therefore, cardiac contractile function. We tested this idea by examining myocardial function in isolated hearts and Ca2+ channel activity in isolated cardiac myocytes. Myocardial function was tested in Langendorff perfused hearts and ICa-L were recorded in the whole-cell patch configuration by applying double pulses from a holding potential of −80 mV and then normalized to the peak amplitudes of control currents. 6-Aminonicotinamide, a competitive inhibitor of G6PD, increased pCO2 and decreased pH. Additionally, 6-aminonicotinamide inhibited G6PD activity, reduced NADPH levels, attenuated peak ICa-L amplitudes, and decreased left ventricular developed pressure and ±dp/dt. Finally, dialyzing NADPH into cells from the patch pipette solution attenuated the suppression of ICa-L by 6-aminonicotinamide. Likewise, in G6PD-deficient mice, G6PD insufficiency in the heart decreased GSH-to-GSSG ratio, superoxide, cholesterol and acetyl CoA. In these mice, M-mode echocardiographic findings showed increased diastolic volume and end-diastolic diameter without changes in the fraction shortening. Taken together, these findings suggest that inhibiting G6PD activity and reducing NADPH levels alters metabolism and leads to inhibition of L-type Ca2+ channel activity. Notably, this pathway may be involved in modulating myocardial contractility under physiological and pathophysiological conditions during which the pentose phosphate pathway-derived NADPH redox is modulated (e.g., ischemia-reperfusion and heart failure).
Highlights
Voltage-gated L-type Ca2+ channels play an important role in the regulation of myocardial contractile function by controlling Ca2+ entry and Ca2+-induced Ca2+ release from sarcoplasmic reticulum in cardiac myocytes
To test that idea and to shed light on the role played by glucose-6-phosphate dehydrogenase (G6PD) and NADPH in regulating L-type Ca2+ channel and heart function, we studied the effects of 6aminonicotinamide (6AN), a competitive G6PD inhibitor [15], and G6PD deficiency on cardiac metabolism and function, and Ltype Ca2+ activity in isolated cardiac myocytes
We found that inhibition of G6PD caused small but significant reduction in metabolism, L-type Ca2+ currents, which are partially reversed by administration of exogenous NADPH, and cardiac function
Summary
Voltage-gated L-type Ca2+ channels play an important role in the regulation of myocardial contractile function by controlling Ca2+ entry and Ca2+-induced Ca2+ release from sarcoplasmic reticulum in cardiac myocytes. Their activity is modulated by a variety of neurotransmitters, hormones and autacoids via regulatory processes involving multiple enzymatic reactions. Myocytes, while testosterone inhibits both native and human recombinant L-type Ca2+ channels from ventricular myocytes, single T-type Ca2+ channels from neonatal rat cardiomyocytes [3,4], and both L- and T-type Ca2+ channels stably expressed in A7r5 and HEK 293 cells [5,6] The effects of both 17b-estradiol and testosterone are voltage-independent. It is known that application of some steroids to cardiac myocytes shifts the current-voltage (I–V) relation and steady-state inactivation curve to more negative potentials, the mechanisms by which steroid hormones inhibit Ca2+ channel activity remain unclear
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