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Abstract: Insulin-like growth factor-1 (IGF-1) plays important roles in the regulation of neuronal development. The electrical activity of Na+ channels is crucial for the regulation of synaptic formation and maintenance/repair of neuronal circuits. Here, we examined the effects of chronic IGF-1 treatment on cell surface expression and function of Na+ channels. In cultured bovine adrenal chromaffin cells expressing NaV1.7 isoform of voltage-dependent Na+ channels, chronic IGF-1 treatment increased cell surface [3H]saxitoxin binding by 31%, without altering the Kd value. In cells treated with IGF-1, veratridine-induced 22Na+ influx, and subsequent 45Ca2+ influx and catecholamine secretion were augmented by 35%, 33%, 31%, respectively. Pharmacological properties of Na+ channels characterized by neurotoxins were similar between nontreated and IGF-1-treated cells. IGF-1-induced up-regulation of [3H]saxitoxin binding was prevented by phosphatydil inositol-3 kinase inhibitors (LY204002 or wortmannin), or Akt inhibitor (Akt inhibitor IV). Glycogen synthase kinase-3 (GSK-3) inhibitors (LiCl, valproic acid, SB216763 or SB415286) also increased cell surface [3H]saxitoxin binding by ∼33%, whereas simultaneous treatment of IGF-1 with GSK-3 inhibitors did not produce additive increasing effect on [3H]saxitoxin binding. IGF-1 (100nM) increased Ser437-phosphorylated Akt and Ser9-phosphorylated GSK-3β, and inhibited GSK-3β activity. Treatment with IGF-1, LiCl or SB216763 increased protein level of Na+ channel α-subunit; it was prevented by cycloheximide. Either treatment increased α-subunit mRNA level by ∼48% and accelerated α-subunit gene transcription by ∼30% without altering α-subunit mRNA stability. Thus, inhibition of GSK-3β caused by IGF-1 up-regulates cell surface expression of functional Na+ channels via acceleration of α-subunit gene transcription. [Copyright &y& Elsevier]

Abstract: In cultured bovine adrenal chromaffin cells expressing NaV1.7 isoform of voltage-dependent Na+ channels, we have previously reported that lithium chloride (LiCl) inhibits function of Na+ channels independent of glycogen synthase kinase-3 (GSK-3) (Yanagita et al., 2007). Here, we further examined the effects of chronic lithium treatment on Na+ channels. LiCl treatment (1–30 mM, ≥12 h) increased cell surface [3H]saxitoxin ([3H]STX) binding by ∼32% without altering the affinity of [3H]STX binding. This increase was prevented by cycloheximide and actinomycin D. SB216763 and SB415286 (GSK-3 inhibitors) also increased cell surface [3H]STX binding by ∼31%. Simultaneous treatment with LiCl and SB216763 or SB415286 did not produce an increased effect on [3H]STX binding compared with either treatment alone. LiCl increased Na+ channel α-subunit mRNA level by 32% at 24 h. LiCl accelerated α-subunit gene transcription by 35% without altering α-subunit mRNA stability. In LiCl-treated cells, LiCl inhibited veratridine-induced 22Na+ influx as in untreated cells. However, washout of LiCl after chronic treatment enhanced veratridine-induced 22Na+ influx, 45Ca2+ influx and catecholamine secretion by ∼30%. Washout of LiCl after 24 h treatment shifted concentration–response curve of veratridine upon 22Na+ influx upward, without altering its EC50 value. Ptychodiscus brevis toxin-3 allosterically enhanced veratridine-induced 22Na+ influx by two-fold in untreated and LiCl-treated cells. Whole-cell patch-clamp analysis indicated that I–V curve and steady-state inactivation/activation curves were comparable between untreated and LiCl-treated cells. Thus, GSK-3 inhibition by LiCl up-regulated cell surface NaV1.7 via acceleration of α-subunit gene transcription, enhancing veratridine-induced Na+ influx, Ca2+ influx and catecholamine secretion. [Copyright &y& Elsevier]

Insulin-like growth factor-1 (IGF-1) plays important roles in the regulation of neuronal development. The electrical activity of Na(+) channels is crucial for the regulation of synaptic formation and maintenance/repair of neuronal circuits. Here, we examined the effects of chronic IGF-1 treatment on cell surface expression and function of Na(+) channels. In cultured bovine adrenal chromaffin cells expressing Na(V)1.7 isoform of voltage-dependent Na(+) channels, chronic IGF-1 treatment increased cell surface [(3)H]saxitoxin binding by 31%, without altering the Kd value. In cells treated with IGF-1, veratridine-induced (22)Na(+) influx, and subsequent (45)Ca(2+) influx and catecholamine secretion were augmented by 35%, 33%, 31%, respectively. Pharmacological properties of Na(+) channels characterized by neurotoxins were similar between nontreated and IGF-1-treated cells. IGF-1-induced up-regulation of [(3)H]saxitoxin binding was prevented by phosphatydil inositol-3 kinase inhibitors (LY204002 or wortmannin), or Akt inhibitor (Akt inhibitor IV). Glycogen synthase kinase-3 (GSK-3) inhibitors (LiCl, valproic acid, SB216763 or SB415286) also increased cell surface [(3)H]saxitoxin binding by ∼ 33%, whereas simultaneous treatment of IGF-1 with GSK-3 inhibitors did not produce additive increasing effect on [(3)H]saxitoxin binding. IGF-1 (100 nM) increased Ser(437)-phosphorylated Akt and Ser(9)-phosphorylated GSK-3β, and inhibited GSK-3β activity. Treatment with IGF-1, LiCl or SB216763 increased protein level of Na(+) channel α-subunit; it was prevented by cycloheximide. Either treatment increased α-subunit mRNA level by ∼ 48% and accelerated α-subunit gene transcription by ∼ 30% without altering α-subunit mRNA stability. Thus, inhibition of GSK-3β caused by IGF-1 up-regulates cell surface expression of functional Na(+) channels via acceleration of α-subunit gene transcription., (Copyright © 2011 Elsevier Ltd. All rights reserved.)