3-4-dihydro-5-(4-(1-piperidinyl)butoxy)-1(2h)-isoquinolinone and benzamide

An excessive activation of poly(ADP-ribose) polymerase (PARP) has been proposed to play a key role in post-ischemic neuronal death. We examined the neuroprotective effects of the PARP inhibitors benzamide, 6(5H)-phenanthridinone, and 3,4-dihydro-5-[4-1(1-piperidinyl)buthoxy]-1(2H)-isoquinolinone in three rodent models of cerebral ischemia. Increasing concentrations of the three PARP inhibitors attenuated neuronal injury induced by 60 min oxygen-glucose deprivation (OGD) in mixed cortical cell cultures, but were unable to reduce CA1 pyramidal cell loss in organotypic hippocampal slices exposed to 30 min OGD or in gerbils following 5 min bilateral carotid occlusion. We then examined the necrotic and apoptotic features of OGD-induced neurodegeneration in cortical cells and hippocampal slices using biochemical and morphological approaches. Cortical cells exposed to OGD released lactate dehydrogenase into the medium and displayed ultrastructural features of necrotic cell death, whereas no caspase-3 activation nor morphological characteristics of apoptosis were observed at any time point after OGD. In contrast, a marked increase in caspase-3 activity was observed in organotypic hippocampal slices after OGD, together with fluorescence and electron microscope evidence of apoptotic neuronal death in the CA1 subregion. Moreover, the caspase inhibitor Z-VAD-FMK reduced OGD-induced CA1 pyramidal cell loss. These findings suggest that PARP overactivation may be an important mechanism leading to post-ischemic neurodegeneration of the necrotic but not of the apoptotic type.

Topics: Animals; Apoptosis; Benzamides; Brain Ischemia; Caspase 3; Caspases; Cell Death; Cell Line; Cerebral Cortex; Enzyme Inhibitors; Gerbillinae; In Vitro Techniques; Isoquinolines; Microscopy, Electron; Microscopy, Fluorescence; Necrosis; Neurons; Neuroprotective Agents; Phenanthrenes; Piperidines; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Pyramidal Cells; Rats; Rats, Wistar

Nitric oxide (NO) and peroxynitrite, formed from NO and superoxide anion, have been implicated as mediators of neuronal damage following focal ischemia, but their molecular targets have not been defined. One candidate pathway is DNA damage leading to activation of the nuclear enzyme, poly(ADP-ribose) polymerase (PARP), which catalyzes attachment of ADP ribose units from NAD to nuclear proteins following DNA damage. Excessive activation of PARP can deplete NAD and ATP, which is consumed in regeneration of NAD, leading to cell death by energy depletion. We show that genetic disruption of PARP provides profound protection against glutamate-NO-mediated ischemic insults in vitro and major decreases in infarct volume after reversible middle cerebral artery occlusion. These results provide compelling evidence for a primary involvement of PARP activation in neuronal damage following focal ischemia and suggest that therapies designed towards inhibiting PARP may provide benefit in the treatment of cerebrovascular disease.

Topics: Adenosine Triphosphate; Animals; Benzamides; Brain; Cells, Cultured; Cerebral Cortex; Cerebrovascular Circulation; DNA Damage; Enzyme Activation; Enzyme Inhibitors; Hemodynamics; Immunity, Innate; Ischemic Attack, Transient; Isoquinolines; Mice; Mice, Knockout; N-Methylaspartate; NAD; Neurons; Neurotoxins; Nitrates; Nitric Oxide; Piperidines; Poly(ADP-ribose) Polymerases