Nitric oxide induces rapid, calcium‐dependent release of vesicular glutamate and ATP from cultured rat astrocytes

Abstract

Nitric oxide (NO; 1 M) or an NO donor (500 M diethylenetriamine-nitric oxide, DETA-NONOate) caused rapid glutamate and ATP release from cultured rat cortical astrocytes. NO-induced glutamate release was prevented by calcium chelators (EGTA or BAPTA-AM) and an inhibitor of vesicular exocytosis (botulinum neurotoxin C, BoTx-C), but not by a glutamate transport inhibitor, L-trans-pyrrolidine-2,4-dicarboxylate (t-PDC), a cyclooxygenase inhibitor (indomethacin), or an inhibitor of soluble guanylate cyclase 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ), and was not induced by mitochondrial respiratory inhibitors (myxothiazol or azide). Similarly to glutamate, NO-induced ATP release was also completely blocked by BAPTA-AM and BoTx-C, suggesting again a vesicular, calcium-dependent mechanism of release. Addition of DETA-NONOate (500 M) to fura-2-loaded astrocytes induced a rapid, transient increase in intracellular calcium levels followed by a lower, sustained level of calcium entry. The latter was blocked by gadolinium (1 M), an inhibitor of capacitative Ca 2ϩ entry. Thus, NO appears to cause rapid exocytosis of vesicular glutamate and ATP from astrocytes by raising intracellular calcium levels. Astrocytes activated by lipopolysaccharide/endotoxin and interferon-␥ to express inducible NO synthase (iNOS) maintained substantially higher extracellular glutamate levels than nonactivated cells or activated cells treated with an iNOS inhibitor (1400W), but the rate of glutamate uptake by these cells was similar. This suggests that NO from inflammatoryactivated astrocytes causes release of astrocytic glutamate. NO-induced release of astrocytic glutamate and ATP may be important in physiological or pathological communication between astrocytes and neurons. GLIA 40:312-323, 2002.

Maxi-anion channel as a candidate pathway for osmosensitive ATP release from mouse astrocytes in primary culture

“…A recent single-vesicle imaging study has provided evidence for astrocyte exocytosis [52]. On the basis of sensitivity to bafilomycin A 1 , tetanus neurotoxin, botulinum neurotoxin C and Ca 2+ chelators, it was suggested that mechanostress-or NO-induced ATP release from rat astrocytes is mediated by exocytosis [23,28]. In the present study, however, intracellular Ca 2+ chelation by pretreatment with a Ca 2+ chelator, BAPTA-AM, and an inhibitor of vesicular transport, BFA, failed to inhibit swelling-induced ATP release from mouse astrocytes.…”

“…Astrocytes have been shown to release ATP in a non-lytic manner in response to osmotic swelling [11], mechanical stimulation [8,[20][21][22][23], deprivation of extracellular Ca 2+ [22][23][24], and stimulation with glutamate [7], UTP [25,26], noradrenaline [27] or NO [28]. However, the precise pathway for ATP release remains controversial.…”

Localization of SNARE proteins and secretory organelle proteins in astrocytes in vitro and in situ

“…Astrocytic release of glutamate (Bezzi et al, 1998;Parpura et al, 1994;Pasti et al, 2001), ATP (Bal-Price et al, 2002;Coco et al, 2003), of the secretory granule protein secretogranin II (Calegari et al, 1999), and of atrial natriuretic peptide (Krzan et al, 2003) were found to depend on an increase in intracellular Ca 2+ concentrations. Furthermore, prolonged incubation of astrocytes with the botulinum neurotoxins A, C 1 and B, or with tetanus toxin (which cleave the SNARE proteins SNAP-25, syntaxin I, and synaptobrevin II/cellubrevin, respectively) resulted in attenuated receptor-evoked glutamate release (Araque et al, 2000;Jeftinija et al, 1997;Pasti et al, 2001).…”

Pannexin 1 in erythrocytes: Function without a gap

“…In particular, ATP is released from erythrocytes, which, under physiological conditions, are vesicle-free (7). Various channels have been implicated in the process, including CFTR (cystic fibrosis transmembrane conductance regulator), connexin 43 (Cx43) hemichannels, a volumeregulated channel (VRAC), and the purinergic receptor P2X7 (5,6,(8)(9)(10)(11). However, the evidence for their involvement falls short because of questionable specificity of the pharmacological blockers used to determine channel identity.…”

“…Two general release modes have been proposed: (i) vesicular release akin to the exocytotic release of transmitters and (ii) channel-mediated release. Although vesicular ATP release is well documented (5,6), it cannot account for all of the ATP release phenomena. In particular, ATP is released from erythrocytes, which, under physiological conditions, are vesicle-free (7).…”