Short- and long-term differential effects of neuroprotective drug NS-7 on voltage-dependent sodium channels in adrenal chromaffin cells.

摘要: In cultured bovine adrenal chromaffin cells, NS-7 [4-(4-fluorophenyl)-2-methyl-6-(5-piperidinopentyloxy) pyrimidine hydrochloride], a newly-synthesized neuroprotective drug, inhibited veratridine-induced 22Na+ influx via voltage-dependent Na+ channels (IC50=11.4 μM). The inhibition by occurred in the presence of ouabain, an inhibitor Na+,K+ ATPase, but disappeared at higher concentration veratridine, and upon washout NS-7. NS-7 attenuated 45Ca2+ Ca2+ (IC50=20.0 μM) catecholamine secretion (IC50=25.8 μM). Chronic (⩾12 h) treatment cells with increased cell surface [3H]-STX binding 86% (EC50=10.5 μM; t1/2=27 h), did not alter KD value; it was prevented cycloheximide, protein synthesis, or brefeldin A, vesicular transport from trans-Golgi network, associated levels channel α- β1-subunit mRNAs. In subjected to chronic treatment, caused veratridine (site 2 toxin), α-scorpion venom 3 toxin) β-scorpion 4 suppressed even after extensive NS-7, remained depressed veratridine; however, either venom, Ptychodiscus brevis toxin-3 5 enhanced as nontreated cells. These results suggest that acute binds site reversibly inhibits channels, thereby reducing gating secretion. Chronic up-regulates translational externalization events, persistently without impairing cooperative interaction between functional domains channels. Keywords: Neuroprotective drugs, sodium up-regulation, [3H]-saxitoxin binding, Northern blot, influx, secretion, cells Introduction It has become increasingly evident noninactivating currents (Taylor, 1993) (Catterall, 1992) may initiate detrimental cascade hypoxia/ischaemia-induced injury, such massive overflow glutamate catecholamines (Toner & Stamford, 1997), intracellular overload reversed operation Na+-Ca2+ exchanger (Obrenovitch Richards, 1995; Urenjak Obrenovitch, 1996), well cytotoxic formation nitric oxide (NO) (Strijbos et al., 1996). A few studies have revealed during hypoxia, steady-state inactivation shifted more hyperpolarizing potential human cortical neurons (Cummins rat hippocampus (O'Reilly presumably compensatory defensive response against hypoxia-induced neuronal injury. Also, density fluctuated brain hypoxia (Perez-Pinzon 1992; Xia Haddad, 1994; 1999). NS-7 hydrochloride] is agent. Previous vivo vitro cerebral cortex shown degradation cytoskeletal fodrin (Takagaki 1997) injury (Tatsumi 1998b) 10–30 μM, cellular mechanisms for neuroprotection are postulated be attributed blockade channels. brain, bound Ki value 1 μM (Shimidzu toxin α-subunit, major subunit forming ion-pore sites 1–5 1992); 10–30 μM diminished dopamine (Itoh 1998) interacts transmembrane segment 6 domain I (IS6) α-subunit (Trainer 1996) activates 1992). depolarization-elicited IC50 7.8 μM NG108-15 (Suma 1997). decreased L-, N-, T-type values 7.3, 4.5, 17.1 μM, respectively cerebrocortical neurons, high K+- activation NO synthase inhibiting L- P/Q-type 1998a; Oka 1999). K+-induced 10 μM 1998), affect release 1997). Na+ α-subunits arise multiple genes their alternative splicing (Dietrich whereas β1-subunits structurally similar among various tissues (Makita 1994). (embryologically derived neural crest), (Yamamoto homologous tetrodotoxin (TTX)/saxitoxin (STX) 1 toxin)-sensitive neuroendocrine type (hNE-Na) (Klugbauer 1995). showed subsequent depolarization increases (Wada 1985a,1985b; Lopez 1995), triggering exocytic (Ito 1980). We found antiepileptic drug valproic acid raised mRNA levels, thus enhancing Similar observations were made long-term anticonvulsant carbamazepine (Yoshimura 1998). phenytoin (Sashihara 1994) antiarrhythmic mexiletine (Kang up-regulation modifying cardiac pathophysiologies. Our present study examined whether/how short- might using (PbTx-3) 1987; Catterall, effects on evaluated.