Treatment of hippocampal neurons with cyclosporin A results in calcium overload and apoptosis which are independent on NMDA receptor activation.
Calcineurin is a ubiquitous calcium/calmodulin dependent protein phosphatase that has been shown to regulate the activity of ion channels, glutamate release, and synaptic plasticity. In the present study we show that CsA, a specific inhibitor of calcineurin, affects the survival of cultures developed from hippocampal dentate gyrus. Mixed neuronal-glial cultures exposed to 8 - 40 microM CsA undergo cell death characterized by apoptotic changes in cellular and nuclear morphology. TUNEL-positive staining was observed only in neurons that developed pyknotic morphology after treatment with 8 microM CsA for 24 - 72 h. Immunocytochemical staining with an anti-GFAP monoclonal antibody revealed that astrocytes from mixed neuronal/glial cultures were unaffected by exposure to CsA at doses toxic for neurons and all TUNEL-positive cells were neurons. MK-801, a noncompetitive inhibitor of glutamate receptor, does not inhibit the appearance of TUNEL-positive neurons and apoptotic changes in nuclear morphology. Preincubation of cells with 8 microM CsA increased basal intracellular calcium level in time dependent manner and decreased relative calcium response to glutamate. Application of 1 microM MK-801 had no effect on CsA-induced changes in Ca(2+) level. Our findings suggest that the neuronal death after CsA treatment is not a result of glutamate excitotoxicity and the increase in intracellular calcium concentration in neurons is not dependent on calcium influx via NMDA channel.
Kaminska
B
B
Laboratory of Transcription Regulation, Nencki Institute of Experimental Biology, 3 Pasteur str., 02-093 Warsaw, Poland. bozenakk@nencki.gov.pl
Figiel
I
I
Pyrzynska
B
B
Czajkowski
R
R
Mosieniak
G
G
D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016572:Cyclosporine / Q000494:pharmacology*; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D020287:In Situ Nick-End Labeling; D009457:Neuroglia / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D016194:Receptors, N-Methyl-D-Aspartate / Q000187:drug effects* / Q000378:metabolism; D013997:Time Factors
D016194:Receptors, N-Methyl-D-Aspartate; D016291:Dizocilpine Maleate; D016572:Cyclosporine; D002118:Calcium