Despite the fact that Ca2+ transport into the sarcoplasmic reticulum (SR) of muscle cells is electrogenic, a potential difference is not maintained across the SR membrane. To achieve electroneutrality, compensatory charge movement must occur during Ca2+ uptake. To examine the role of Cl- in this charge movement in smooth muscle cells, Ca2+ transport into the SR of saponin-permeabilized smooth muscle cells was measured in the presence of various Cl- channel blockers or when I-, Br-, or SO42- was substituted for Cl-. Calcium uptake was inhibited in a dose-dependent manner by 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) and by indanyloxyacetic acid 94 (R(+)-IAA-94), but not by niflumic acid or 4,4'-dinitrostilbene-2,2'-disulfonic acid (DNDS). Smooth muscle SR Ca2+ uptake was also partially inhibited by the substitution of SO42- for Cl-, but not when Cl- was replaced by I- or Br-. Neither NPPB nor R(+)-IAA-94 inhibited Ca2+ uptake into cardiac muscle SR vesicles at concentrations that maximally inhibited uptake in smooth muscle cells. These results indicate that Cl- movement is important for charge compensation in smooth muscle cells and that the Cl- channel or channels involved are different in smooth and cardiac muscle cells.

Ca2+ transients in isolated cardiac ventricular myocytes and the amount of Ca2+ that could be released from the sarcoplasmic reticulum (SR) in these cells by caffeine were reduced in the presence of tamoxifen. To examine the effects of tamoxifen on the cardiac muscle SR directly, isolated SR vesicles and fluorimetry methods were used to measure the uptake of Ca2+ by the SR and the ATPase activity of the SR Ca2+ pump. SR Ca2+ uptake was inhibited by tamoxifen at concentrations greater than 2.4 microM. Half-maximal inhibition was seen at approximately 5 microM. Inhibition of uptake was not due to the development of a substantial tamoxifen-dependent leak of Ca2+ from the SR or to a direct inhibitory effect of tamoxifen on the ATPase activity of the SR Ca2+ pump. In addition to its effect on SR Ca2+ uptake, tamoxifen also reduced the rate at which stored Ca2+ could be released from the SR by the Ca2+ ionophore 4-bromo A23187. Our results are consistent with the hypothesis that tamoxifen inhibits an ion current that accompanies Ca2+ movement across the SR membrane. This possibility is also consistent with the known inhibitory action of tamoxifen on some types of Cl- and K+ channels.

Neural activity is important for establishing proper connectivity in the developing visual system. Tetrodotoxin blockade of sodium (Na(+))-dependent action potentials impairs the refining of synaptic connections made by developing retinal ganglion cells (RGCs), but does not affect their ability to get out to their target. Although this may suggest neural activity is not required for the directed extension of RGC axons, in many species developing RGCs express additional, Na(+)-independent ionic mechanisms. To test whether the ability of RGC axons to extend in a directed fashion is influenced by membrane excitability, we blocked the principal modulators of the neural activity of a neuron, voltage-dependent potassium (Kv) channels. First, we showed that RGCs and their growth cones express Kv channels when they are growing through the brain on the way to their main midbrain target, the optic tectum. Second, a Kv channel blocker, 4-aminopyridine (4-AP), was applied to the developing Xenopus optic projection. Blocking Kv channels inhibited RGC axon extension and caused aberrant routing of many RGC fibers. With the higher doses, <25% of embryos had a normal optic projection. These data suggest that Kv channel activity regulates the guidance of growing axons in the vertebrate brain.

Accumulating evidence suggests that voltage-dependent potassium (Kv) channels have important and varied roles in the development of neuronal and non-neuronal cell types. They have been implicated in processes such as proliferation, cell adhesion, migration, neurite outgrowth, and axon guidance. In this study, we used antibodies against several electrically active Kv channel alpha-subunits (Kv1-4) to describe the spatial and temporal expression patterns of Kv channel subunits in Xenopus laevis retinal ganglion cell (RGC) somata, axons, and growth cones. We found that RGCs express Kv1.3-, Kv1.5-, Kv3.4-, and Kv4.2-like subunits. Each subunit displayed unique cellular and subcellular distributions. Moreover, the expression patterns changed considerably over the major period of Xenopus retinal cell genesis and differentiation. Weak or no immunoreactivity was observed with antibodies against Kv1.1, Kv1.2, Kv1.4, Kv1.6, and Kv3.2 subunits in RGCs or other retinal cell types. In support of our previous pharmacologic evidence implicating Kv channels in RGC axon outgrowth, we found that Kv1.5-, Kv3.4-, and Kv4.2-like proteins, but not Kv1.3-like subunits, are abundantly expressed in RGC growth cones.