Calciseptine isolated from the black mamba toxin selectively blocks cardiac Cav1.2 (alpha1C) versus Cav1.3 (alpha1D) L-type calcium channels

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Introduction: Selective blockers are important tools for identifying the physiological role of ion channels isoforms. L-type calcium channels are high voltage activated channels sensitive to dihydropyridine (DHP). Two distinct isoforms (1C/Cav1.2 and 1D/Cav1.3) mediate calcium entry into cardiac cells to trigger respectively contraction and heart rate. Many other classes of drugs have been shown to target L-type calcium channels but poor selectivity has been reported between Cav1.2 and Cav1.3. Aim: Our goal was to identify a pharmacological tool able to discriminate between between Cav1.2 and Cav1.3. isoforms. We focused on calciseptine (Cas) , a snake toxin purified from mamba venom reported to decrease vascular and cardiac contraction via a L-type calcium channel inhibition but without affecting heart rate (De Weille et al, 2001). Material and methods: The contraction amplitude and heart rate were investigated in Langendorf perfused hearts from both wild-type and mice carrying Cav1.2 channels insensitive to DHP. Whole cell patch clamp experiments were used to study the effect of Cas on L-type calcium currents in ventricular cells, sino-atrial cells known to express both isoforms and recombinant Cav1.2 and Cav1.3 currents in HEK cells. In addition, the effect of Cas was investigated on HVA neuronal calcium channel subtypes(N,P/Q), T-type current and hERG potassium current.Results: Calciseptine strongly decreased the amplitude of contractions (around 80% at 100 nM) while no modification of the heart rate was observed up to 1uM Cas. The frequency of spontaneous AP recorded from sinus node using the patch clamp technique was also unaffected by 100nM Cas by contrast with the effect of 3 µM nifedipine, which in isolated mouse heart , decreased both contraction and heart rate, induced arrhythmias and strongly reduced spontaneous AP frequency in isolated sinus node. Voltage-clamp experiments in ventricular and sinusal cells and recombinant Cav1.2 and Cav1.3 currents confirmed the selective inhibition of Cas for the Cav 1.2 isoform and the lack of effect on other calcium channel subtypes.Conclusions: Our data evidence that calciseptine acts as a specific blocker of Cav1.2 L-type calcium current and could be an important tool to study the physiological and pathophysiological role of Cav1.2 versus Cav1.3 in heart but in various cell types including nendocrine cells and neurons where they are co-expressed .

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