Poly(ADP-ribose) polymerase: early involvement in glutamate-induced neurotoxicity in cultured cerebellar granule cells.
Glutamate neurotoxicity is correlated with an increase of cytosolic free Ca2+. In some cell systems, activation of Ca2+ dependent endonucleases or formation of free radicals can damage DNA and activate the chromatin bound enzyme poly(ADP-ribose) polymerase (pADPRP). We have investigated whether pADPRP may be involved in glutamate neurotoxicity in vitro. Cerebellar granule cells at 12 days in culture when treated with a toxic dose of glutamate (100 microM) showed a rapid and transient increase of polyADP-ribose immunoreactivity. Cellular immunostaining was heterogeneous and returned to control levels after washout of glutamate. In the same cell preparations glutamate elicited a marked increase in enzyme protein immunoreactivity which persisted at later times. Non-toxic doses of glutamate did not affect immunostaining. In another set of experiments, pADPRP mRNA was increased 30 min after glutamate. In order to investigate the role of pADPRP in glutamate-mediated neurotoxicity, structurally different inhibitors of pADPRP (3-aminobenzamide, benzamide,3-aminophthalhydrazide) and their inactive analogues (benzoic acid and phthalimide) were tested in this model. Addition of the inhibitors to cultures 60 min before and during the 30 min of glutamate treatment prevented neuronal death by 60-100%, assessed 24 hr later. Glutamate-induced Ca2+ influx was not affected. Inactive analogues failed to afford neuroprotection. These data indicate that not only is pADPRP activated by the early, possibly Ca(2+)-mediated mechanisms initiated by glutamate, but that it might also actively contribute to the subsequent neuronal death.
Cosi
C
C
Fidia Research Laboratories, Abano Terme, Italy.
Suzuki
H
H
Milani
D
D
Facci
L
L
Menegazzi
M
M
Vantini
G
G
Kanai
Y
Y
Skaper
S D
SD
D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D001565:Benzoates / Q000494:pharmacology; D019817:Benzoic Acid; D002118:Calcium / Q000502:physiology; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D002525:Cerebellar Cortex / Q000166:cytology*; D004790:Enzyme Induction / Q000187:drug effects; D018698:Glutamic Acid / Q000633:toxicity*; D008165:Luminol / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D009474:Neurons / Q000187:drug effects* / Q000201:enzymology; D010797:Phthalimides / Q000494:pharmacology; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley
D001549:Benzamides; D001565:Benzoates; D009419:Nerve Tissue Proteins; D010797:Phthalimides; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D012333:RNA, Messenger; C037431:phthalimide; D018698:Glutamic Acid; D008165:Luminol; C037689:benzamide; C025160:3-aminobenzamide; D019817:Benzoic Acid; D011065:Poly(ADP-ribose) Polymerases; D002118:Calcium