Longitudinal distribution of Na⁺ and Ca²⁺ channels and β‐adrenoceptors on the sarcolemmal membrane of frog cardiomyocytes

Abstract

The distribution of L‐type Ca2+ and tetrodotoxin‐sensitive Na+ channels and of β‐adrenergic receptors was examined in frog ventricular myocytes using the whole‐cell patch‐clamp technique and a double capillary for extracellular microperfusion. Rod‐shaped cells (250–300 μm long) were sealed at both ends to two patch‐clamp pipettes and positioned transversally at different positions between the mouths of two micro‐capillaries separated by a thin wall. A combination of nifedipine (1 μm) and tetrodotoxin (0.3 μm) (blocking solution) was added to one capillary in order to inhibit macroscopic Ca2+ and Na+ currents (ICa and INa, respectively) in the part of the cell exposed to this capillary. Moving the cell in 10–20 μm steps from the control capillary to the capillary containing the blocking solution induced step decreases in ICa and INa amplitudes. Complete block of both currents occurred when the entire cell was exposed to the blocking solution. Each step decrease in current was due to the loss of activity of the functional Ca2+ and Na+ channels present in the slice of sarcolemmal membrane newly exposed to the blocking solution. These step current changes allowed longitudinal mapping of current density for Ca2+ and Na+ channels on the sarcolemmal membrane. Addition of a submaximal concentration of isoprenaline (10 nm) to the control capillary induced a local increase in ICa which enabled examination of the distribution of functional β‐adrenergic receptors as well. Our results demonstrate that Ca2+ and Na+ channels and β‐adrenergic receptors are equally and essentially uniformly distributed on the sarcolemmal membrane of frog ventricular myocytes.

β2‐Adrenergic receptor overexpression in the developing mouse heart: evidence for targeted modulation of ion channels

“…Although this is the first report of cAMP compartmentalization in cardiac cells in a manner that distinguishes target ion channel regulation by β 2 ‐AR overexpression, our results complement those of others who have focused on calcium entry and calcium transients in cardiac myocytes using pharmacological approaches (Xiao et al 1994; Jurevicius & Fischmeister, 1996, 1997; Zhou et al 1997). Previous studies have linked β 2 ‐AR‐induced changes in contraction, cytosolic Ca 2+ , and L‐type Ca 2+ channel currents to activation of adenylyl cyclase and subsequent increases in intracellular cAMP (Skeberdis et al 1997).…”

Role of cyclic nucleotide phosphodiesterase isoforms in cAMP compartmentation following beta2-adrenergic stimulation of ICa,L in frog ventricular myocytes

“…The ‘local’ I Ca,L corresponds to the amount of current that disappeared after withdrawing Ca 2+ from S2 (in this case 35 pA). Since L‐type Ca 2+ channels are uniformly distributed along the cell length (Jurevicius & Fischmeister, 1997), distant and local I Ca,L amplitudes give an indication of how much cell membrane is exposed to each solution. In all experiments, the cells were positioned in such a way that distant and local I Ca,L did not differ by more than 15 %, so that it could be assumed that a similar amount of cell membrane was exposed to each solution.…”

Local response of L‐type Ca2+ current to nitric oxide in frog ventricular myocytes

“…Thus, perfusion of nominally Ca 2+ ‐free solution as S1 (or S2) allows spatial separation of I Ca . Since the VDCCs are distributed uniformly in frog cardiomyocytes (Jurevicius & Fischmeister, 1997), I Ca values measured in S1 or S2 are a direct measure of the membrane under the influence of S1 and S2.…”