. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. Figure 3. Changes of intracellular pH as a function of extracellular Na+ concentration. pressed (Fig. 2). Raising the extracellular tonicity of the normal Na+ medium by sucrose to the same level of the high Na+ medium did not suppress AChR clustering. Thus, this effect could not be attributed to an increase in osmotic pressure. This suppression due to high Na+ could be reversed by 1 mM amilonde, which inhibits the Na+-H+ exchange in the plasma membrane (Aronson, 1985). Furthermore, application of 10-20 mA/NH4Cl, a weak base
. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. Figure 3. Changes of intracellular pH as a function of extracellular Na+ concentration. pressed (Fig. 2). Raising the extracellular tonicity of the normal Na+ medium by sucrose to the same level of the high Na+ medium did not suppress AChR clustering. Thus, this effect could not be attributed to an increase in osmotic pressure. This suppression due to high Na+ could be reversed by 1 mM amilonde, which inhibits the Na+-H+ exchange in the plasma membrane (Aronson, 1985). Furthermore, application of 10-20 mA/NH4Cl, a weak base which causes rapid elevation of cytoplasmic pH, suppressed the bead-induced formation of AChR clusters. These results suggest that the intracellular pH may be involved in the regulation of bead-induced clus- ter formation. This was tested by intracellular pH mea- surements. When the extracellular Na:+ was elevated from 60 mM to 100 mM there was a cytoplasmic alka- linization of pH units over a 45 min period; this could be suppressed by 1 mM amiloride (Fig. 3). Bath application of polycations, such as polylysine or polyor- nithine, also effected a large alkalinization of the cyto- plasm (Fig. 4). Discussion These studies show that an increase in intracellular Ca2+ is a necessary condition for the induction of AChR clustering induced by polycation-coated beads. Depriva- tion of extracellular Ca2+ also inhibits the formation of AChR clusters induced by nerve (Henderson et ai. 1984), by molecules extracted from the basement mem- brane of nicotinic synapses (Wallace, 1988), or by con- stant electric field (Stollberg and Fraser. 1988). Thus, all these stimuli may cause an opening of the Ca:+ channels in the plasma membrane. The calcium influx resulting from this action may trigger the process of AChR cluster- ing. Our previous studies have also shown that calmodu- lin may mediate the bead-induced process in Xenopus muscle cells (Peng, 1984). Recent evidence shows that the basemen
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