Sialic acid and the surface charge associated with hyperpolarization-activated, inward rectifying channels

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Abstract
The whole-cell configuration of the patch-clamp technique was used with cultured pacemaker cells from the rabbit sinoatrial node to test the hypothesis that sialic acid residues (NANA) constitute much of the negative surface charge associated with hyperpolarization-activated, inward rectifying channels. Activation-voltage relationships (between −70 and −140 mV) were determined for hyperpolarization-activated (inward rectifying) current (i f). Addition of 10mm Ca2+ shifted the half-activation potential (V 1/2) from −89.5±0.9 mV to −77.9±2.6 mV (PV 1/2 from −86.8±1.4 mV to −75.0±1.7 mV (P2+ reduced the amplitude ofi f significantly. Incubation of cells with a highly purified preparation of neuraminidase (0.1–2.0 U/ml, 1 hr, 37°C), an enzyme that selectively removes NANA from glycoproteins and glycolipids, failed to alterV 1/2 or the amplitude ofi f significantly. Pretreatment of cells with neuraminidase (1.0 U/ml, 1 hr, 37°C) failed to alter the positive shift ofV 1/2 produced by dimethonium. The results suggest that NANA does not constitute the negative surface charge associated with hyperpolarization-activated, inward rectifying channels.