Inhibition of multiple pathways accounts for the antiapoptotic effects of flavopiridol on potassium withdrawal-induced apoptosis in neurons.
Serum and potassium (S/K) deprivation is a well-known apoptotic model in cerebellar granule neurons (CGNs), used to study the efficacy of potential neuroprotective drugs. The objective of this study was to determine the pathways involved in the neuroprotective role of flavopiridol, a pan-inhibitor of cyclin-dependent kinases (CDKs), upon S/K withdrawal-induced apoptosis in CGNs. Cell death in primary cultures of rat CGNs was accompanied by chromatin condensation and activation of caspases-3, -6, and -9. Caspase-3 activity was also evaluated by cleavage of 120-kDa alpha-spectrin. Flavopiridol (1 microM) prevented caspase activation and abolished apoptotic features mediated by S/K withdrawal. Re-entry in the cell cycle is also involved in apoptotic neuronal cell death. Flavopiridol (1 microM) inhibited DNA synthesis as measured by BrdU incorporation, thus enhancing proliferating cell nuclear antigen expression. Serum/potassium (S/K) deprivation induced apoptotic cell death mediated by the activation of several kinases such as glycogen synthase kinase-3beta and CDK5, as well as the breakdown of p35 in the neurotoxic fragment p25; inactivation of myocyte enhancer factor-2 (MEF2) was also found. Pretreatment with flavopiridol prevented these biochemical and molecular alterations. Taken together, these findings suggest an apoptotic route in CGNs after S/K withdrawal mediated by the activation of several kinases involved in cell cycle deregulation and MEF2 inactivation. We propose that the antiapoptotic properties of flavopiridol are mediated through kinase pathway inhibition.
Verdaguer
Ester
E
Unitat de Farmacologia i Farmacognòsia, Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, E-08028 Barcelona, Spain.
Jordà
Elvira G
EG
Alvira
Daniel
D
Jiménez
Andrés
A
Canudas
Anna Maria
AM
Folch
Jaume
J
Rimbau
Victor
V
Pallàs
Mercè
M
Camins
Antoni
A
D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002453:Cell Cycle / Q000187:drug effects; D002531:Cerebellum / Q000502:physiology*; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors; D005419:Flavonoids / Q000494:pharmacology*; D005434:Flow Cytometry; D008959:Models, Neurological; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D010880:Piperidines / Q000494:pharmacology*; D011188:Potassium / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley
D005419:Flavonoids; D010880:Piperidines; D047428:Protein Kinase Inhibitors; C077990:alvocidib; D018844:Cyclin-Dependent Kinases; D011188:Potassium