title: Activation of brain nitric oxide synthase in depolarized human temporal cortex slices: differential role of voltage-sensitive calcium channels.
abstract: 1. Nitric oxide (NO) synthase activity was studied in slices of human temporal cortex samples obtained in neurosurgery by measuring the conversion of L-[3H]-arginine to L-[3H]-citrulline. 2. Elevation of extracellular K+ to 20, 35 or 60 mM concentration-dependently augmented L-[3H]-citrulline production. The response to 35 mM KCl was abolished by N(G)-nitro-L-arginine (100 microM) demonstrating NO synthase specific conversion of L-arginine to L-citrulline. Increasing extracellular MgCl2 concentration up to 10 mM also prevented the K+ (35 mM)-induced NO synthase activation, suggesting the absolute requirement of external calcium ions for enzyme activity. 3. However, the effect of high K+ (35 mM) on citrulline synthesis was insensitive to the antagonists of ionotropic and metabotropic glutamate receptors dizocilpine (MK-801), 6-nitro-7-sulphamoylbenzo(f)-quinoxaline-2-3-dione (NBQX) or L-2-amino-3-phosphonopropionic acid (L-AP3) as well as to the nicotinic receptor antagonist, mecamylamine. 4. The 35 mM K+ response was insensitive to omega-conotoxin GVIA (1 microM) and nifedipine (100 microM), but could be prevented in part by omega-agatoxin IVA (0.1 and 1 microM). The inhibition caused by 0.1 microM omega-agatoxin IVA (approximately 30%) was enhanced by adding omega-conotoxin GVIA (1 microM) or nifedipine (100 microM). Further inhibition (up to above 70%) could be observed when the three Ca2+ channel blockers were added together. Similarly, synthetic FTX 3.3 arginine polyamine (sFTX) prevented (50% at 100 microM) the K+-evoked NO synthase activation. This effect of sFTX was further enhanced (up to 70%) by adding 1 microM omega-conotoxin GVIA plus 100 microM nifedipine. No further inhibition could be observed upon addition of MK-801 or/and NBQX. 5. It was concluded that elevation of extracellular [K+] causes NO synthase activation by external Ca2+ entering cells mainly through channels of the P/Q-type. Other Ca2+ channels (L- and N-type) appear to contribute when P/Q-channels are blocked.
authors.affiliation: Istituto di Farmacologia e Tossicologia, Università di Genova, Italy.
authors.forename: G
authors.initials: G
authors.lastname: Fontana
authors.forename: E
authors.initials: E
authors.lastname: Fedele
authors.forename: M
authors.initials: M
authors.lastname: Cossu
authors.forename: C
authors.initials: C
authors.lastname: Munari
authors.forename: M
authors.initials: M
authors.lastname: Raiteri
mesh_terms: D000409:Alanine / Q000031:analogs & derivatives / Q000494:pharmacology; D001120:Arginine / Q000032:analysis / Q000737:chemistry; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000502:physiology*; D002956:Citrulline / Q000096:biosynthesis; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D008464:Mecamylamine / Q000494:pharmacology; D018733:Nicotinic Antagonists / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019335:Nitroarginine / Q000494:pharmacology; D011188:Potassium / Q000378:metabolism; D011189:Potassium Chloride / Q000494:pharmacology; D011810:Quinoxalines / Q000494:pharmacology; D013702:Temporal Lobe / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology
chemical_list: D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004791:Enzyme Inhibitors; D018691:Excitatory Amino Acid Antagonists; D018733:Nicotinic Antagonists; D011810:Quinoxalines; C000211:2-amino-3-phosphonopropionic acid; C062865:2,3-dioxo-6-nitro-7-sulfamoylbenzo(f)quinoxaline; D019335:Nitroarginine; D002956:Citrulline; D011189:Potassium Chloride; D008464:Mecamylamine; D016291:Dizocilpine Maleate; D001120:Arginine; D019001:Nitric Oxide Synthase; D000409:Alanine; D011188:Potassium