Selective induction of brain type II Na+ channels by nerve growth factor.

Abstract

Cells derived from a rat pheochromocytoma (PC12 cells) can generate an action potential only upon treatment with nerve growth factor. Using electrophysiological methods, we found that the appearance of action potentials in nerve growth factor-treated PC12 cells can be explained by an increase in the density of Na' channels. The functional properties of Na' channels in PC12 cells are similar to those described for peripheral nerves but appear to be different from Na' channels synthesized in Xenopus oocytes injected with brain type II Na'-channel mRNA. To determine if PC12 cells express the brain type II Na+ -channel gene, we performed RNase-protection analyses using probes that can distinguish between the brain type I and type II Na'-channel mRNAs. The results from these studies indicate that undifferentiated PC12 cells express the type II but not the type I Na+ -channel gene. Treatment with nerve growth factor increases expression of the type II Na+-channel gene but has no effect on type I gene expression. Our findings suggest that N+ -channel excitability in PC12 cells is due to the specific induction of the brain type II gene by nerve growth factor.Neurons can be distinguished from inexcitable types of cells by their capacity to generate an action potential. The appearance of excitability occurs early in neuronal differentiation (1-3) and results from the insertion of voltage-dependent ion channels into the membrane (4). PC12 cells, derived from a tumor of adrenal tissue (5), have proven useful for studying the factors that regulate excitability. Treatment with nerve growth factor (NGF), for example, enables PC12 cells to generate action potentials. Electrophysiological studies have indicated that the action potential in NGF-treated PC12 cells is due to activation of voltage-dependent Na' channels (6, 7). It has also been shown that the number of saxitoxin binding sites on PC12 cells increases after treatment with NGF (8). Therefore, even though voltage-dependent K+ and Ca2" channels also appear during differentiation of PC12 cells (9, 10), it is the expression of Na+ channels that best accounts for the development of the action potential. The effect of NGF on the density of functional Na+ channels has not been examined in detail. Previous electrophysiological studies in PC12 cells have been limited either to measurement of action potentials (6, 7) or to single-channel Na' currents (11) and have not provided quantitative estimates of the density of functional Na+ channels. Therefore, in this study we utilized the whole-cell voltage-clamp technique in order to quantitate the effect of NGF on Na+-channel density. These experiments were performed in concert with molecular studies designed to measure changes in Na+-channel gene expression in response to treatment with NGF.Molecular studies have indicated that excitable cells express different Na'-channel mRNAs (12)(13)(14). cDNA clones coding for two structurally distinct voltage-dependent Na' channels in rat brain (termed type I and type II) have been ide...

Analysis of Mutant Platelet-Derived Growth Factor Receptors Expressed in PC12 Cells Identifies Signals Governing Sodium Channel Induction during Neuronal Differentiation

“…A probe specific for GAPDH mRNA was also included in the hybridization as an internal control for variations in harvesting and handling the RNA samples during analysis (see Materials and Methods). Similar to wild-type PC12 cells (18,49), in the sublines expressing wild-type human PDGF receptors, there were low levels of type II ␣-subunit mRNA in the absence of growth factor treatment and easily discernible increases (3.4 Ϯ 0.43-fold; n ϭ 3) in response to NGF (Fig. 1C).…”

“…The numerous morphological and biochemical changes that occur include neurite outgrowth, the induction of immediate-early genes such as c-fos, and an increase in the expression of neuronspecific genes, including those encoding for the metalloproteinase transin or the pore-forming ␣ subunit of the voltagedependent sodium (Na ϩ ) channel (for reviews, see references 25 and 26). Of the seven Na ϩ channel ␣-subunit genes known to be expressed in the nervous system (1,12,21,56,65,72), wild-type PC12 cells express the PN-1 and type II genes (12,49). NGF causes a rapidly induced, transient increase in PN-1…”

Bradykinin inhibits a potassium M‐like current in rat pheochromocytoma PC12 cells

“…NGF increases both the density of several receptors [14,24], and the expression of type II sodium channels [25], but not of high-voltage-activated calcium currents [26]. To assess if NGF altered the Mcurrent density on PC12 cells, we compared the current normalized for capacitance in control and in cells treated with NGF for more than 4 days.…”

Role of neurotrophin signalling in the differentiation of neurons from dorsal root ganglia and sympathetic ganglia

“…In frog sympathetic neurons, calcium conductances are sensitive to NGF levels in vivo and the regulation of calcium and sodium currents has been demonstrated in vitro. In rat PC12 cells, sodium currents and various sodium channel transcripts are induced by NGF (Mandel et al 1988;D'Arcangelo et al 1993).…”