Volume regulated anion channel currents of rat hippocampal neurons and their contribution to oxygen-and-glucose deprivation induced neuronal death.

Volume-regulated anion channels (VRAC) are widely expressed chloride channels that are critical for the cell volume regulation. In the mammalian central nervous system, the physiological expression of neuronal VRAC and its role in cerebral ischemia are issues largely unknown. We show that hypoosmotic medium induce an outwardly rectifying chloride conductance in CA1 pyramidal neurons in rat hippocampal slices.

Functions of astrocytes and their potential as therapeutic targets.

Astrocytes are often referred to, and historically have been regarded as, support cells of the mammalian CNS. Work over the last decade suggests otherwise-that astrocytes may in fact play a more active role in higher neural processing than previously recognized. Because astrocytes can potentially serve as novel therapeutic targets, it is critical to understand how astrocytes execute their diverse supportive tasks while maintaining neuronal health.

Functions of mature mammalian astrocytes: a current view.

Before the roles of normal, mature astrocytes in the mammalian CNS can be discussed, we first need to define these cells. A definition proposed here is that such a class is best defined as consisting of the protoplasmic and fibrous astrocytes of the gray and white matter, respectively, the Bergmann glia of the molecular layer of the cerebellum, and the Muller cells of the retina. It is concluded that the established properties and functions of these mature astrocytes are essential support for neuronal activity, in the sense of Claude Bernard's principle of maintaining "la fixité du milieu intérieur.

Electrical coupling of astrocytes in rat hippocampal slices under physiological and simulated ischemic conditions.

Mammalian protoplasmic astrocytes are extensively coupled through gap junction channels but the biophysical properties of these channels under physiological and ischemic conditions in situ are not well defined. Using confocal morphometric analysis of biocytin-filled astrocytic syncytia in rat hippocampal CA1 stratum radiatum we found that each astrocyte directly couples, on average, to 11 other astrocytes with a mean interastrocytic distance of 45 microm. Voltage-independent and bidirectional transjunctional currents were always measured between directly coupled astrocyte pairs in dual voltage-clamp recordings, but never from astrocyte-NG2 glia or astrocyte-interneuron pairs.

TWIK-1 and TREK-1 are potassium channels contributing significantly to astrocyte passive conductance in rat hippocampal slices.

Expression of a linear current-voltage (I-V) relationship (passive) K(+) membrane conductance is a hallmark of mature hippocampal astrocytes. However, the molecular identifications of the K(+) channels underlying this passive conductance remain unknown. We provide the following evidence supporting significant contribution of the two-pore domain K(+) channel (K(2P)) isoforms, TWIK-1 and TREK-1, to this conductance.

Tamoxifen mediated estrogen receptor activation protects against early impairment of hippocampal neuron excitability in an oxygen/glucose deprivation brain slice ischemia model.

Pretreatment of ovarectomized rats with estrogen shows long-term protection via activation of the estrogen receptor (ER). However, it remains unknown whether activation of the ER can provide protection against early neuronal damage when given acutely. We simulated ischemic conditions by applying oxygen and glucose deprived (OGD) solution to acute male rat hippocampal slices and examined the neuronal electrophysiological changes.

Two distinct modes of hypoosmotic medium-induced release of excitatory amino acids and taurine in the rat brain in vivo.

A variety of physiological and pathological factors induce cellular swelling in the brain. Changes in cell volume activate several types of ion channels, which mediate the release of inorganic and organic osmolytes and allow for compensatory cell volume decrease. Volume-regulated anion channels (VRAC) are thought to be responsible for the release of some of organic osmolytes, including the excitatory neurotransmitters glutamate and aspartate.

Supportive or information-processing functions of the mature protoplasmic astrocyte in the mammalian CNS? A critical appraisal.

It has been proposed that astrocytes should no longer be viewed purely as support cells for neurons, such as providing a constant environment and metabolic substrates, but that they should also be viewed as being involved in affecting synaptic activity in an active way and, therefore, an integral part of the information-processing properties of the brain. This essay discusses the possible differences between a support and an instructive role, and concludes that any distinction has to be blurred. In view of this, and a brief overview of the nature of the data, the new evidence seems insufficient to conclude that the physiological roles of mature astrocytes go beyond a general support role.

Tamoxifen as a powerful neuroprotectant in experimental stroke and implications for human stroke therapy.

Several recent studies from the author's laboratory have shown that tamoxifen, at higher concentrations than used for breast cancer and given i.v., can substantially prevent tissue infarct and behavioral deficits in reversible and permanent rat focal stroke models for up to at least 14 days after initiation of ischemia.

Resveratrol attenuates early pyramidal neuron excitability impairment and death in acute rat hippocampal slices caused by oxygen-glucose deprivation.

Accumulating evidence indicates that the polyphenol resveratrol (trans-3, 5, 4"-trihydroxystibene, RVT) potently protects against cerebral ischemia neuronal damage due to its oxygen free radicals scavenging and antioxidant properties. However, it is unknown whether RVT can attenuate ischemia-induced early impairment of neuronal excitability. To address this question, we simulated ischemic conditions by applying oxygen-glucose deprivation (OGD) to acute rat hippocampal slices and examined the effect of RVT on OGD-induced pyramidal neuron excitability impairment using whole-cell patch clamp recording.

Neuroprotection by alpha 2-adrenergic agonists in cerebral ischemia.

Ischemic brain injury is implicated in the pathophysiology of stroke and brain trauma, which are among the top killers worldwide, and intensive studies have been performed to reduce neural cell death after cerebral ischemia. Alpha 2-adrenergic agonists have been shown to improve the histomorphological and neurological outcome after cerebral ischemic injury when administered during ischemia, and recent studies have provided considerable evidence that alpha 2-adrenergic agonists can protect the brain from ischemia/reperfusion injury. Thus, alpha 2-adrenergic agonists are promising potential drugs in preventing cerebral ischemic injury, but the mechanisms by which alpha 2-adrenergic agonists exert their neuroprotective effect are unclear.

DCPIB, a specific inhibitor of volume regulated anion channels (VRACs), reduces infarct size in MCAo and the release of glutamate in the ischemic cortical penumbra.

Previous studies have indicated that volume regulated anion channels (VRACs) may be involved in the pathology of the ischemic brain cortical penumbra due to activation of VRAC-mediated excitatory amino-acid (EAA) release. To assess this we had studied neuroprotection and EAA release inhibition by a potent VRAC inhibitor, tamoxifen. However, tamoxifen inhibits several other neurodamaging processes.

Oxygen and glucose deprivation-induced changes in astrocyte membrane potential and their underlying mechanisms in acute rat hippocampal slices.

Accumulating evidence indicates a significant astrocytic involvement in cerebral ischemia neuropathology, but little is known about the immediate astrocytic responses to ischemia insults in terms of electrophysiology and their pathologic implications. We show that astrocytes in acute rat hippocampal slices responded reversibly to more than 30 mins oxygen and glucose deprivation (OGD) treatment with depolarized membrane potentials (V(m)) in whole-cell current clamp recording. This depolarization was multiphasic, showing an initial approximately 11 mins small-amplitude depolarization plateau, followed by a 6-mins accelerated depolarization, and then a second plateau.

Neuroprotection by tamoxifen in focal cerebral ischemia is not mediated by an agonist action at estrogen receptors but is associated with antioxidant activity.

We have previously shown that tamoxifen can induce marked neuroprotection after middle cerebral artery occlusion (MCAo) in rats and have described two possible mechanisms of action: namely, inhibition of EAA release and inhibition of nNOS activity. In this study we tested other potential mechanisms. Namely, agonist action at estrogen receptors and an antioxidative action.

Anion channels in astrocytes: biophysics, pharmacology, and function.

The chloride/anion channels that have been so far identified in cultured astrocytes and those that have been confirmed in situ by a combination of mRNA identification, immunocytochemistry, and biophysical studies are reviewed. It is emphasized that we are just beginning to describe such channels and analyze their functions in astrocytes. The best-studied anion channels studied so far are those known as volume-regulated anion channels (VRACs).

Development of gap junctions in hippocampal astrocytes: evidence that whole cell electrophysiological phenotype is an intrinsic property of the individual cell.

Gap junction communication between astrocytes is prevalent and has been proposed to be involved in several astrocyte functions. One such proposal involves gap junctions in potassium spatial buffering. However, little is known about the developmental time course of gap junction coupling and how much the syncytium affects whole cell measurements of ion currents.

Neuroprotection by pyrroloquinoline quinone (PQQ) in reversible middle cerebral artery occlusion in the adult rat.

Pyrroloquinoline quinone (PQQ) is a naturally occurring redox cofactor that acts as an essential nutrient, antioxidant, and redox modulator. It has previously been reported to reduce infarct size in 7-day-old rat pups with an in vivo cerebral hypoxia/ischemia model (Jensen et al., 1994).

Pharmacological comparison of swelling-activated excitatory amino acid release and Cl- currents in cultured rat astrocytes.

Ubiquitously expressed volume-regulated anion channels (VRACs) are chloride channels which are permeable to a variety of small organic anions, including the excitatory amino acids (EAAs) glutamate and aspartate. Broad spectrum anion channel blockers strongly reduce EAA release in cerebral ischaemia and other pathological states associated with prominent astrocytic swelling. However, it is uncertain whether VRAC serves as a major pathway for EAA release from swollen cells.

Development of GLAST(+) astrocytes and NG2(+) glia in rat hippocampus CA1: mature astrocytes are electrophysiologically passive.

Glia show marked heterogeneity in terms of electrophysiology in the developing brain, and two major types can be identified based on GFAP or NG2 expression. However, it remains to be determined if such an electrophysiological diversity holds for the adult brain and how GFAP and NG2 lineage glia are associated with different electrophysiological phenotypes during the course of development. To address these fundamental questions, we performed in situ whole cell recording from morphologically identified glia from the rat hippocampal CA1 region from postnatal (P) days 1-106 and double-stained postrecorded cells with GLAST and NG2 antibodies.

Behavioral and histological neuroprotection by tamoxifen after reversible focal cerebral ischemia.

We have previously shown that tamoxifen can reduce infarct sizes measured by 2,3,5,-triphenyltetrazolium chloride (TTC) staining at 72 h after 2 h of reversible middle cerebral artery occlusion (rMCAo) in rats. In this study, we tested whether improvement is found in both behavioral measures of protection and by histological measures of infarcted tissue at 7 and 14 days after 2 h rMCAo. Tamoxifen (10 mg/kg) was given once by intravenous injection 1 h after reperfusion, i.

Astrocytic swelling in cerebral ischemia as a possible cause of injury and target for therapy.

In this viewpoint article, I summarize data showing that the astrocytic swelling that occurs early after the acute CNS pathologies ischemia and traumatic brain injury is damaging. We have proposed that one reason may be the release of excitatory amino acids (EAA) via volume-activated anion channels (VRACs) that are activated by such swelling. This release could be a target for therapy, which could involve blocking the astrocytic swelling or the release mechanisms.

Hydrogen peroxide potentiates volume-sensitive excitatory amino acid release via a mechanism involving Ca2+/calmodulin-dependent protein kinase II.

Excessive excitatory amino acid (EAA) release in cerebral ischemia is a major mechanism responsible for neuronal damage and death. A substantial fraction of ischemic EAA release occurs via volume-regulated anion channels (VRACs). Hydrogen peroxide (H2O2), which is abundantly produced during ischemia and reperfusion, activates a number of protein kinases critical for VRAC functioning and has recently been reported to activate VRACs.

ATP regulates anion channel-mediated organic osmolyte release from cultured rat astrocytes via multiple Ca2+-sensitive mechanisms.

Ubiquitously expressed volume-regulated anion channels (VRACs) are activated in response to cell swelling but may also show limited activity in nonswollen cells. VRACs are permeable to inorganic anions and small organic osmolytes, including the amino acids aspartate, glutamate, and taurine. Several recent reports have demonstrated that neurotransmitters or hormones, such as ATP and vasopressin, induce or strongly potentiate astrocytic whole cell Cl(-) currents and amino acid release, which are inhibited by VRAC blockers.