Acetylcholine-induced, calcium-dependent norepinephrine outflow from peripheral human lymphocytes.
Catecholamines (CA) were studied in peripheral human lymphocytes, as well as in the supernatants, after incubation with L-tyrosine and L-dihydroxyphenylalanine (L-Dopa) for 1 h. The effect that the addition of acetylcholine (ACh), Veratridine, lonomycin or KCI had on the outflow of norepinephrine (NE) from lymphocytes was also studied. The effect of the addition of methoxyverapamil (D600, a Ca2+ channel blocker) and cholinergic antagonists had on the ACh-induced NE outflow was assessed. CA were determined by HPLC-ECD, both in the supernatant and in the cell lysates. L-Tyrosine and L-Dopa significantly (P < 0.01) increased intracellular NE. Neither L-tyrosine, L-Dopa, nor vehicle induced a detectable outflow of NE to the supernatants. ACh [120 microM], Veratridine [100 microM], Ionomycin [10 microM] and KCl [50 mM] (with or without the simultaneous addition of L-tyrosine or L-Dopa) all induced a detectable outflow of NE to the supernatant when added 5 min before the end of incubation. NE was not detectable in the supernatant when the chemicals were added 10 to 20 min before the end of the incubation. When the chemicals were added at lower concentrations, erratic secretion or no secretion whatsoever was observed. D600 [100 microM] was able to significantly (P < 0.01) reduce the ACh-induced NE outflow. Tetraethylammonium (nicotinic antagonist), but not atropine (muscarinic antagonist), significantly (P < 0.001) decreased the ACh-induced NE outflow. The outflow of NE from peripheral human lymphocytes was seen. NE secretion seems to be ACh- and calcium-dependent since Veratridine, Ionomycin and KCl are able to induce Ca2+ entry by means of various mechanisms. The Ca2+ channel blocker employed in this study (D600) reduced the ACh-dependent NE outflow. We can conclude that both ACh (through nicotinic receptors) and calcium are involved in the outflow of NE from peripheral human lymphocytes.
Department of Internal Medicine, San Martino Hospital, Genoa, Italy. nrmusso@unige.it
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Musso
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Brenci
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Indiveri
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Lotti
D000109:Acetylcholine / Q000494:pharmacology*; D000328:Adult; D002118:Calcium / Q000502:physiology*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005260:Female; D005711:Gallopamil / Q000494:pharmacology; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007980:Levodopa / Q000494:pharmacology; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D009638:Norepinephrine / Q000378:metabolism*; D014443:Tyrosine / Q000494:pharmacology; D014701:Veratridine / Q000494:pharmacology
D002121:Calcium Channel Blockers; D005711:Gallopamil; D014443:Tyrosine; D007980:Levodopa; D015759:Ionomycin; D014701:Veratridine; D000109:Acetylcholine; D002118:Calcium; D009638:Norepinephrine