The effects of calbindin D-28K and parvalbumin antisense oligonucleotides on the survival of cultured Purkinje cells.
The role of calcium binding proteins, calbindin D-28k (CaB) and parvalbumin (PV) in Purkinje cell survival was investigated using oligonucleotide antisense strategy. Purkinje cell enriched cultures were prepared from the cerebella of 0-1 day old Balb/c mouse pups. Purkinje cells were identified by size, asymmetric arbors, immunoreactivity to CaB and PV, uptake of gamma-aminobutyric acid (GABA) and failure to express glial fibrillary acidic protein. The cells at different days in vitro were treated with antisense or mismatched antisense phosphorothioate oligonucleotides for CaB and PV mRNA (complexed with lipofectin). Neuronal specific [3H]-GABA uptake was used as a measure of Purkinje cell survival. The cultures treated for 24 h with antisense oligos (CaB+PV) showed a significant decrease in [3H]-GABA uptake as compared with the cultures treated with lipofectin alone or with lipofectin + mismatched antisense oligos to CaB and PV mRNA. The results of the present study suggest that the expression of calcium buffering proteins CaB and PV may have a significant involvement in Purkinje cell viability.
Vig
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PJ
Department of Neurology, University of Mississippi Medical Center, Jackson 39216, USA. pvig@neurology.umsmed.edu
McDaniel
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D000818:Animals; D000831:Animals, Newborn; D000911:Antibodies, Monoclonal; D064026:Calbindins; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016650:Fluorescein-5-isothiocyanate; D005455:Fluorescent Antibody Technique; D051379:Mice; D008807:Mice, Inbred BALB C; D016501:Neurites / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D010320:Parvalbumins / Q000037:antagonists & inhibitors* / Q000502:physiology; D011689:Purkinje Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D064030:S100 Calcium Binding Protein G / Q000037:antagonists & inhibitors* / Q000502:physiology; D013997:Time Factors; D014404:Tubulin / Q000378:metabolism; D005680:gamma-Aminobutyric Acid / Q000493:pharmacokinetics
D000911:Antibodies, Monoclonal; D064026:Calbindins; D016376:Oligonucleotides, Antisense; D010320:Parvalbumins; D064030:S100 Calcium Binding Protein G; D014404:Tubulin; D005680:gamma-Aminobutyric Acid; D016650:Fluorescein-5-isothiocyanate