Nitric oxide (NO) and zinc (Zn2+) are implicated in the pathogenesis of cerebral ischemia and neurodegenerative diseases. However, their relationship and the molecular mechanism of their neurotoxic effects remain unclear. Here we show that addition of exogenous NO or NMDA (to increase endogenous NO) leads to peroxynitrite (ONOO-) formation and consequent Zn2+ release from intracellular stores in cerebrocortical neurons. Free Zn2+ in turn induces respiratory block, mitochondrial permeability transition (mPT), cytochrome c release, generation of reactive oxygen species (ROS), and p38 MAP kinase activation. This pathway leads to caspase-independent K+ efflux with cell volume loss and apoptotic-like death. Moreover, Zn2+ chelators, ROS scavengers, Bcl-xL, dominant-interfering p38, or K+ channel blockers all attenuate NO-induced K+ efflux, cell volume loss, and neuronal apoptosis. Thus, these data establish a new form of crosstalk between NO and Zn2+ apoptotic signal transduction pathways that may contribute to neurodegeneration.

The conceptual understanding of hippocampal function has been challenged recently by the finding that new granule cells are born throughout life in the mammalian dentate gyrus (DG). The number of newborn neurons is dynamically regulated by a variety of factors. Kainic acid-induced seizures, a rodent model of human temporal lobe epilepsy, strongly induce the proliferation of DG neurogenic progenitor cells and are also associated with long-term cognitive impairment. We show here that the antiepileptic drug valproic acid (VPA) potently blocked seizure-induced neurogenesis, an effect that appeared to be mainly mediated by inhibiting histone deacetylases (HDAC) and normalizing HDAC-dependent gene expression within the epileptic dentate area. Strikingly, the inhibition of aberrant neurogenesis protected the animals from seizure-induced cognitive impairment in a hippocampus-dependent learning task. We propose that seizure-generated granule cells have the potential to interfere with hippocampal function and contribute to cognitive impairment caused by epileptic activity within the hippocampal circuitry. Furthermore, our data indicate that the effectiveness of VPA as an antiepileptic drug may be partially explained by the HDAC-dependent inhibition of aberrant neurogenesis induced by seizure activity within the adult hippocampus.

Myocyte enhancer factor-2 (MEF2) transcription factors are activated by p38 mitogen-activated protein kinase during neuronal and myogenic differentiation. Recent work has shown that stimulation of this pathway is antiapoptotic during development but proapoptotic in mature neurons exposed to excitotoxic or other stress. We now report that excitotoxic (N-methyl-D-aspartate) insults to mature cerebrocortical neurons activate caspase-3, -7, in turn cleaving MEF2A, C, and D isoforms. MEF2 cleavage fragments containing a truncated transactivation domain but preserved DNAbinding domain block MEF2 transcriptional activity via dominant interference. Transfection of constitutively active MEF2 (MEF2C-CA) rescues MEF2 transcriptional activity after N-methyl-D-aspartate insult and prevents neuronal apoptosis. Conversely, dominant-interfering MEF2 abrogates neuroprotection by MEF2C-CA. These results define a pathway to excitotoxic neuronal stress͞apo-ptosis via caspase-catalyzed cleavage of MEF2. Additionally, we show that similar MEF2 cleavage fragments are generated in vivo during focal stroke damage. Hence, this pathway appears to have pathophysiological relevance in vivo. Paradoxically, we and others have shown that activation of the p38 mitogen-activated protein kinase (MAPK) pathway can lead to either antiapoptotic or proapoptotic effects on neurons (1-4). An important transcription factor that is activated by the p38 stress kinase pathway and thought to mediate these life and death events, at least in part, is myocyte enhancer factor-2 (MEF2) (5, 6). There are four MEF2 proteins (MEF2A, -B, -C, and -D). They represent transcription factors in the MADS (MCM1-agamous-deficiens-serum response factor) family, signifying that they contain a MADS box and act in concert with other MADS domain factors as well as other classes of transcription factors (7-10). Previously, we had cloned and characterized MEF2C and shown that it was the predominant MEF2 family member in cerebrocortical neurons (7), with lesser contributions of MEF2A and -D in this neuronal population (11, 12). Olson and colleagues have shown that MEF2 proteins are also highly expressed in cardiac myocytes and induce myogenesis in conjunction with basic helix-loop-helix transcription factors, as judged from both in vitro and in vivo models; other groups have reported concordant findings (13-15). Moreover, recent work from our group and others has shown that in vitro transfection of stem-like P19 cells or murine embryonic stem cells with a constitutively active form of MEF2C (MEF2C-CA) was antiapoptotic and induced a phenotype with both muscle and neuronal characteristics (4, 16). Forced expression of a dominantinterfering form of MEF2C (MEF2C-DN) prevented retinoicacid-induced neurogenesis of these cells. Additionally, when constitutively active MEF2A was transfected into postmitotic cerebellar granule cell neurons in the face of an insult, such as extracellular K ϩ withdrawal, the MEF2 activity promoted survival (i.e., was antiapoptotic) (17).On the other hand, in...

New neurons are continuously added throughout life to the dentate gyrus of the mammalian hippocampus. During embryonic and early postnatal development, the dentate gyrus is formed in an outside-in layering pattern that may extend through adulthood. In this work we aimed to systematically quantify the relative position of dentate granule cells generated at different ages. We used 5’-bromo-2’-deoxyuridine (BrdU) and retroviral methodologies to birth-date cells born in the embryonic, early postnatal and adult hippocampus and assessed their final position in the adult mouse granule cell layer. We also quantified both developmental and adult-born cohorts of neural progenitor cells that contribute to the pool of adult progenitor cells. Our data confirm that the outside-in layering of the dentate gyrus continues through adulthood and that early-born cells constitute most of the adult dentate gyrus. We also found that a substantial fraction of the dividing cells in the adult dentate gyrus were derived from early-dividing cells and retained BrdU, suggesting that a subpopulation of hippocampal progenitors divides infrequently from early development on.