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We addressed the question of whether glial cells in intact white matter tracts express neurotransmitter receptors and we used Ca+2 signalling as a probe to detect the receptor activation. Corpus callosum slices from postnatal mice were bulk-loaded with the Ca+2-sensitive fluorescent dye fluo-3, and confocal microscopy was used to measure Ca+2 transients in response to neuroligands. Glial cell bodies were intensely dye-loaded and could be discriminated from the diffuse fluorescence of axons. Subpopulations of glial cells from slices obtained at postnatal days 3 to 7 responded with Ca+2 signals to ATP, glutamate, histamine, GABA, norepinephrine, serotonin, angiotensin II, bradykinin, and substance P. These subpopulations showed a distinct overlap; cells which were responsive to substance P always showed Ca+2 signalling in response to histamine, ATP, GABA and high K+ (membrane depolarization). GABA-responsive cells almost always showed a [Ca+2]i increase after membrane depolarization. In brain slices from postnatal day 11 to 18 animals, the Ca+2 responses were evident for glutamate, ATP, and norepinephrine, while GABA, substance P, serotonin, histamine, or angiotensin II rarely elicited a response. This study demonstrates that white matter glial cells in slices exhibit a large repertoire of neurotransmitter responses linked to Ca+2 signalling and that these receptor systems are differentially distributed on sub-populations of glial cells. more...

Schwann cells freshly isolated from the sciatic nerves of neonatal rats respond to exogenously applied ATP with a rapid increase in cytosolic calcium. This increase in [Ca2+]i is mediated by a P2Y-purinergic pathway (Lyons et al.: J. Neurochem. 63:552-560, 1994) and was measured using the calcium indicator dye, fura-2/AM, and a video-enhanced calcium imaging system. The ability to respond to ATP with increases in intracellular calcium is lost over a period of several days in culture; this loss can be prevented or reversed by application of cAMP analogs in a defined medium. We now demonstrate that the direct contact of Schwann cells with neurons also induces and stabilizes this ATP responsiveness. The induction of ATP responsiveness was observed among all Schwann cells contacting neurites, including those forming myelin, and regardless of whether the source of neurons was dorsal root ganglion neurons or superior cervical ganglion neurons. Approximately 85% of Schwann cells responded to ATP over the time studied (72 d in coculture). Addition of axolemma to Schwann cell cultures did not induce ATP responsiveness. We also examined the ATP responsiveness of Schwann cells in situ (excised nerves) using laser-scanning confocal microscopy and the calcium indicator dye, fluo-3/AM. Schwann cells in intact sciatic nerve segments isolated from neonatal and 16-day-old rats exhibited ATP-mediated [Ca2+]i increases. We conclude that neuronal contact is necessary for the expression of the ATP-mediated calcium responses in Schwann cells and that these responses are independent of myelin formation or maintenance. more...

Schwann cells establish close contact with axons during development, and this is maintained throughout life. Signaling by neurotransmitters may play an important role in Schwann cell-axon interaction. Schwann cells were examined for the presence of neuroligland receptors that are linked to increases in levels of cytoplasmic calcium. Schwann cell cultures were prepared from neonatal rat sciatic nerve and, after 0.25, 1, 4, 7, and 14 days in vitro (DIV), loaded with the calcium indicator dye fura 2-AM. The influence of neuroligands on the cytosolic free calcium concentration ([Ca2+]i) was then examined at each time point using a video-based imaging system. Approximately 80-95% of all freshly isolated Schwann cells responded to 10 microM ATP with a three-fold rise in [Ca2+]i. Bradykinin, glutamate, and histamine had no or only partial and inconsistent responses. The ATP-induced calcium response disappeared within 4 DIV. Culturing cells in the presence of cyclic AMP (cAMP) analogues (which induce proliferation and differentiation in vitro) restored the ability of Schwann cells to respond to ATP with increased [Ca2+]i. In the presence of cAMP analogues the extent of recovery of ATP responsiveness was dependent on serum concentration. Fifty to ninety percent of cells regained calcium responsiveness to ATP when grown in medium containing cAMP analogues and 1% serum. These cells also exhibited immunoreactivity to P0 antibody, characteristic of the myelinating lineage. In contrast, only 15-30% of the Schwann cells regained calcium responsiveness when grown in medium containing cAMP analogues and 10% serum.(ABSTRACT TRUNCATED AT 250 WORDS) more...