PubMed Health⌕ Search

Biomedical subjects

Brian A MacVicar

Publications and source records attributed to Brian A MacVicar.

13 recordsLinked to original sources
PubMed2006

p38 mitogen-activated protein kinase contributes to adenosine A1 receptor-mediated synaptic depression in area CA1 of the rat hippocampus.

Adenosine is arguably the most potent and widespread presynaptic modulator in the CNS, yet adenosine receptor signal transduction pathways remain unresolved. Here, we demonstrate a novel mechanism in which adenosine A1 receptor stimulation leads to p38 mitogen-activated protein kinase (MAPK) activation and contributes to the inhibition of synaptic transmission. Western blot analysis indicated that selective A1 receptor activation [with N6-cyclopentyladenosine (CPA)] resulted in rapid increases in phosphorylated p38 (phospho-p38) MAPK immunoreactivity in membrane fractions, and decreases in phospho-p38 MAPK in cytosolic fractions. Immunoprecipitation with a phospho-p38 MAPK antibody revealed constitutive association of this phosphoprotein with adenosine A1 receptors. Phospho-p38 MAPK activation by A1 receptor stimulation induced translocation of PP2a (protein phosphatase 2a) to the membrane. We then examined the actions of p38 MAPK activation in A1 receptor-mediated synaptic inhibition. Excitatory postsynaptic field potentials evoked in area CA1 of the rat hippocampus markedly decreased in response to adenosine (10 microM), the A1 receptor agonist CPA (40 nM), or a 5 min exposure to hypoxia. These inhibitory responses were mediated by A1 receptor activation because the selective antagonist DPCPX (8-cyclopentyl-1,3-dipropylxanthine) (100 nM) prevented them. In agreement with the biochemical analysis, the selective p38 MAPK inhibitor SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole] (25 microM) blocked the inhibitory actions of A1 receptor activation, whereas both the inactive analog SB202474 [4-ethyl-2-(p-methoxyphenyl)-5-(4'-pyridyl)-1H-imidazole] (25 microM) and the ERK 1/2 (extracellular signal-regulated kinase 1/2) MAPK inhibitor PD98059 [2'-amino-3'-methoxyflavone] (50 microM) were ineffective. In contrast, the p38 MAPK inhibitors did not inhibit GABA(B)-mediated synaptic depression. These data suggest A1 receptor-mediated p38 MAPK activation is a crucial step underlying the presynaptic inhibitory effect of adenosine on CA3-CA1 synaptic transmission.

Animals↗
PubMed2006

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). These, as for most channels, have been mainly studied in cultured astrocytes, but some correlative studies have been done in situ, because these channels have been emphasized as release routes for transmitters; namely, excitatory amino acids and ATP. They are activated by cell shape changes and cell swelling, and the release of amino acids and ATP and chloride currents, measured by whole cell clamping, by these processes has been well described, as is also their activation by low concentrations of extracellular ATP. However, the identity of these channels in astrocytes, as in all other cells, remains elusive. The potential involvement of VRACs in pathological states such as stroke, metastasis, and spreading depression is also discussed.

Adenosine Triphosphate↗
PubMed2006

VRACs CARVe a path for novel mechanisms of communication in the CNS.

Because the brain is encased by the skull, the ability to control cell volume in the brain is crucial, and pathological conditions that disturb cell volume homeostasis may severely compromise neural function and survival. Astrocytes are the main cell type to show swelling in response to pathological conditions. More recently, a role for swelling-induced neurotransmitter release from astrocytes under nonpathological conditions has been reported. Astrocytes express a volume-regulated anion channel (VRAC) that is involved in volume homeostasis. In addition to transporting chloride, VRACs allow the efflux of chloride and amino acids such as taurine, glutamate, and aspartate. Glutamate and aspartate are potent activators of neuronal glutamate receptors. Therefore, this nonsynaptic form of cellular communication may modulate neuronal excitability and synaptic activity.

Adenosine Triphosphate↗
PubMed2006

Muscarinic enhancement of R-type calcium currents in hippocampal CA1 pyramidal neurons.

The "toxin-resistant" R-type Ca2+ channels are expressed widely in the CNS and distributed mainly in apical dendrites and spines. They play important roles in regulating signal transduction and intrinsic properties of neurons, but the modulation of these channels in the mammalian CNS has not been studied. In this study we used whole-cell patch-clamp recordings and found that muscarinic activation enhances R-type, but does not affect T-type, Ca2+ currents in hippocampal CA1 pyramidal neurons after N, P/Q, and L-type Ca2+ currents selectively were blocked. M1/M3 cholinergic receptors mediated the muscarinic stimulation of R-type Ca2+ channels. The signaling pathway underlying the R-type enhancement was independent of intracellular [Ca2+] changes and required the activation of a Ca(2+)-independent PKC pathway. Furthermore, we found that the enhancement of R-type Ca2+ currents resulted in the de novo appearance of Ca2+ spikes and in remarkable changes in the firing pattern of R-type Ca2+ spikes, which could fire repetitively in the theta frequency. Therefore, muscarinic enhancement of R-type Ca2+ channels could play an important role in modifying the dendritic response to synaptic inputs and in the intrinsic resonance properties of neurons.

Animals↗
PubMed2006

Ischemia opens neuronal gap junction hemichannels.

Neuronal excitotoxicity during stroke is caused by activation of unidentified large-conductance channels, leading to swelling and calcium dysregulation. We show that ischemic-like conditions [O(2)/glucose deprivation (OGD)] open hemichannels, or half gap junctions, in neurons. Hemichannel opening was indicated by a large linear current and flux across the membrane of small fluorescent molecules. Single-channel openings of hemichannels (530 picosiemens) were observed in OGD. Both the current and dye flux were blocked by inhibitors of hemichannels. Therefore, hemichannel opening contributes to the profound ionic dysregulation during stroke and may be a ubiquitous component of ischemic neuronal death.

Adenosine Triphosphate↗
PubMed2005

Topiramate inhibits the initiation of plateau potentials in CA1 neurons by depressing R-type calcium channels.

PURPOSE: Cholinergic-dependent plateau potentials (PPs) are intrinsically generated conductances that can elicit ictal-type seizure activity. The aim of this study was to investigate the actions of topiramate (TPM) on the generation of PPs. METHODS: We used whole-cell patch-clamp recordings from CA1 pyramidal neurons in rat hippocampal slices to examine the effects of TPM on the PPs. RESULTS: In current-clamp mode, action potentials evoked PPs after cholinergic receptor stimulation. Therapeutically relevant concentrations of TPM (50 microM) depressed the PPs evoked by action potentials. Surprisingly, in voltage-clamp mode, we discovered that the cyclic nucleotide-gated (CNG) current that underlies PP generation (denoted as I(tail)) was not depressed. However, significantly longer depolarizing voltage steps were required to elicit I(tail). This suggested that the calcium entry trigger for evoking PPs was depressed by TPM and not I(tail) itself. TPM had no effect on calcium spikes in control conditions; however, TPM did reduce calcium spikes after cholinergic-receptor stimulation. We recently found that R-type calcium spikes are enhanced by cholinergic-receptor stimulation. Therefore we isolated R-type calcium spikes with a cocktail containing tetrodotoxin, omega-conotoxin MVIIC, omega-conotoxin-GVIA, omega-agatoxin IVA, and nifedipine. R-type calcium spikes were significantly depressed by TPM. We also examined the effects of TPM on recombinant Ca(V)2.3 calcium channels expressed in tsA-201 cells. TPM depressed currents mediated by Ca(V)2.3 subunits by a hyperpolarizing shift in steady-state inactivation. CONCLUSIONS: We have found that TPM reduces ictal-like activity in CA1 hippocampal neurons through a novel inhibitory action of R-type calcium channels.

Action Potentials↗
PubMed2004

Calcium transients in astrocyte endfeet cause cerebrovascular constrictions.

Cerebral blood flow (CBF) is coupled to neuronal activity and is imaged in vivo to map brain activation. CBF is also modified by afferent projection fibres that release vasoactive neurotransmitters in the perivascular region, principally on the astrocyte endfeet that outline cerebral blood vessels. However, the role of astrocytes in the regulation of cerebrovascular tone remains uncertain. Here we determine the impact of intracellular Ca(2+) concentrations ([Ca(2+)](i)) in astrocytes on the diameter of small arterioles by using two-photon Ca(2+) uncaging to increase [Ca(2+)](i). Vascular constrictions occurred when Ca(2+) waves evoked by uncaging propagated into the astrocyte endfeet and caused large increases in [Ca(2+)](i). The vasoactive neurotransmitter noradrenaline increased [Ca(2+)](i) in the astrocyte endfeet, the peak of which preceded the onset of arteriole constriction. Depressing increases in astrocyte [Ca(2+)](i) with BAPTA inhibited the vascular constrictions in noradrenaline. We find that constrictions induced in the cerebrovasculature by increased [Ca(2+)](i) in astrocyte endfeet are generated through the phospholipase A(2)-arachidonic acid pathway and 20-hydroxyeicosatetraenoic acid production. Vasoconstriction by astrocytes is a previously unknown mechanism for the regulation of CBF.

Animals↗
PubMed2003

ATP released from astrocytes during swelling activates chloride channels.

ATP release from astrocytes contributes to calcium ([Ca(2+)]) wave propagation and may modulate neuronal excitability. In epithelial cells and hepatocytes, cell swelling causes ATP release, which leads to the activation of a volume-sensitive Cl(-) current (I(Cl,swell)) through an autocrine pathway involving purinergic receptors. Astrocyte swelling is counterbalanced by a regulatory volume decrease, involving efflux of metabolites and activation of I(Cl,swell) and K(+) currents. We used whole cell patch-clamp recordings in cultured astrocytes to investigate the autocrine role of ATP in the activation of I(Cl,swell) by hypo-osmotic solution (HOS). Apyrase, an ATP/ADP nucleotidase, inhibited HOS-activated I(Cl,swell), whereas ATP and the P2Y agonists, ADPbetaS and ADP, induced Cl(-) currents similar to I(Cl,swell). Neither the P2U agonist, UTP nor the P2X agonist, alpha,beta-methylene ATP, were effective. BzATP was less effective than ATP, suggesting that P2X7 receptors were not involved. P2 purinergic antagonists, suramin, RB2, and pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) reversibly inhibited activation of I(Cl,swell), suggesting that ATP-activated P2Y1 receptors. Thus ATP release mediates I(Cl,swell) in astrocytes through the activation of P2Y1-like receptors. The multidrug resistance protein (MRP) transport inhibitors probenicid, indomethacin, and MK-571 all potently inhibited I(Cl.swell). ATP release from astrocytes in HOS was observed directly using luciferin-luciferase and MK-571 reversibly depressed this HOS-induced ATP efflux. We conclude that ATP release via MRP and subsequent autocrine activation of purinergic receptors contributes to the activation of I(Cl,swell) in astrocytes by HOS-induced swelling.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2002

Activation of presynaptic P2X7-like receptors depresses mossy fiber-CA3 synaptic transmission through p38 mitogen-activated protein kinase.

P2X(7) receptor subunits form homomeric ATP-gated, calcium-permeable cation channels. In this study, we used Western blots and immunocytochemistry to demonstrate that P2X(7) receptors are abundant on presynaptic terminals of mossy fiber synapses in the rat hippocampus. P2X(7)-immunoreactive protein was detected using a specific P2X(7) antibody in Western blots of protein isolated from whole hippocampus and from a subcellular fraction containing mossy fiber synaptosomes. P2X(7) immunoreactivity was colocalized with syntaxin 1A/B-immunoreactivity in mossy fiber terminals in the dentate hilus and stratum lucidum of CA3. Extracellular and whole-cell voltage-clamp recordings in CA3 revealed that bath application of the potent P2X(7) agonist 2',3'-O-(4-benzoylbenzoyl)-ATP (Bz-ATP) caused a long-lasting inhibition of neurotransmission at mossy fiber-CA3 synapses. Consistent with a presynaptic action at mossy fiber synapses, Bz-ATP had no significant effect on neurotransmission at associational-commissural synapses in CA3 but increased paired-pulse facilitation during depression of mossy fiber evoked currents. In addition, Bz-ATP had no postsynaptic effect on holding current or conductance of CA3 neurons. Bz-ATP-induced mossy fiber synaptic depression was blocked by the P2X(7) antagonist oxidized ATP but not by the P2X(1-3,5,6) antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid or the P2Y antagonist reactive blue 2. Finally, an antagonist of p38 MAP kinase activation [4-(4-fluorophenyl)2-(4-methylsulfinylphenyl)5-(4-pyridyl)imidazole] but not extracellular signal-regulated kinase 1/2 MAP kinase (2'-amino-3'-methoxyflavone) blocked the synaptic depression mediated by Bz-ATP, suggesting that this presynaptic inhibition was mediated by activation of p38 MAP kinase. The results of the present study demonstrate that activation of presynaptic P2X(7) receptors depresses mossy fiber-CA3 synaptic transmission through activation of p38 MAP kinase.

Adenosine↗
PubMed2002

Development of Ca2+ hotspots between Lymnaea neurons during synaptogenesis.

Calcium (Ca2+) channel clustering at specific presynaptic sites is a hallmark of mature synapses. However, the spatial distribution patterns of Ca2+ channels at newly formed synapses have not yet been demonstrated. Similarly, it is unclear whether Ca2+ 'hotspots' often observed at the presynaptic sites are indeed target cell contact specific and represent a specialized mechanism by which Ca2+ channels are targeted to select synaptic sites. Utilizing both soma-soma paired (synapsed) and single neurons from the mollusk Lymnaea, we have tested the hypothesis that differential gradients of voltage-dependent Ca2+ signals develop in presynaptic neuron at its contact point with the postsynaptic neuron; and that these Ca2+ hotspots are target cell contact specific. Fura-2 imaging, or two-photon laser scanning microscopy of Calcium Green, was coupled with electrophysiological techniques to demonstrate that voltage-induced Ca2+ gradients (hotspots) develop in the presynaptic cell at its contact point with the postsynaptic neuron, but not in unpaired single cells. The incidence of Ca2+ hotspots coincided with the appearance of synaptic transmission between the paired cells, and these gradients were target cell contact specific. In contrast, the voltage-induced Ca2+ signal in unpaired neurons was uniformly distributed throughout the somata; a similar pattern of Ca2+ gradient was observed in the presynaptic neuron when it was soma-soma paired with a non-synaptic partner cell. Moreover, voltage clamp recording techniques, in conjunction with a fast, optical differential perfusion system, were used to demonstrate that the total whole-cell Ca2+ (or Ba2+) current density in single and paired cells was not significantly different. However, the amplitude of Ba2+ current was significantly higher in the presynaptic cell at its contact side with the postsynaptic neurons, compared with non-contacted regions. In summary, this study demonstrates that voltage-induced Ca2+ hotspots develop in the presynaptic cell, concomitant with the appearance of synaptic transmission between the soma-soma paired cells. The appearance of Ca2+ gradients in presynaptic neurons is target cell contact specific and is probably due to a spatial redistribution of existing channels during synaptogenesis.

Animals↗
PubMed2002

Intrinsic optical signals in the rat optic nerve: role for K(+) uptake via NKCC1 and swelling of astrocytes.

Measurements of extracellular space volume and imaging of intrinsic optical signals (IOSs) have shown that neuronal activity increases light transmittance by causing cellular swelling. However, the cellular mechanisms underlying these volume changes and the contribution of astrocyte swelling to the changes in tissue volume are unclear. In this study, we have investigated IOSs in optic nerves to analyze the mechanisms contributing to these signals in a system consisting of only axons and glial cells. We examined both intact optic nerves and enucleated optic nerves, which contained no axons and consisted primarily of astrocytes. Electrical stimulation of intact optic nerves evoked an increase in light transmittance, which was graded with increasing stimulation frequency and was mimicked by raising extracellular K(+) concentration ([K(+)](o)). The stimulation-induced IOS grew in amplitude and had a time course similar to extracellular space shrinkage. Tetrodotoxin (TTX) blocked the electrically induced but not the high K(+)-induced IOS. In enucleated nerves, light transmittance progressively increased in higher [K(+)](o). The high [K(+)](o)-induced IOSs were reversibly depressed by furosemide and bumetanide, antagonists for Na-K-2Cl cotransport, but were unaltered by TTX. We also used a monoclonal antibody to the NKCC1 form of the Na-K-2Cl cotransporter to show that NKCC1 is expressed in optic nerves as shown in Western blotting and is colocalized in GFAP immunopositive astrocytes. In summary, these results indicated that KCl uptake into astrocytes via an Na-K-2Cl cotransporter during raised [K(+)](o) contributes to the generation of cellular swelling and the intrinsic optical signals.

Action Potentials↗
PubMed2002

Nitric oxide promotes intracellular calcium release from mitochondria in striatal neurons.

Overproduction of nitric oxide by NMDA receptor stimulation is implicated in calcium deregulation and neurodegeneration of striatal neurons. We investigated the involvement of nitric oxide (NO) in inducing intracellular calcium release and in modifying calcium transients evoked by NMDA. NO application (4-10 microM) reversibly and repeatedly increased the intracellular calcium concentration [Ca2+]i in Fura-2- or fluo-3-loaded cultured mouse striatal neurons. NO-induced [Ca2+]i responses persisted in the absence of extracellular calcium, indicating that Ca2+ was released from intracellular stores. The source of calcium was distinct from [Ca2+]i-activated (ruthenium red and ryanodine sensitive) or IP3-activated (thapsigargin-sensitive) Ca2+ stores and was not dependent on cGMP production because a cell permeant analog, 8-bromo-cGMP, did not increase basal [Ca2+]i. Glucose removal potentiated the NO-induced release of [Ca2+]i. In contrast, pretreatment with either the mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone or cyclosporin A, a blocker of the mitochondrial permeability transition pore, prevented the [Ca2+]i increase after NO. The rise in [Ca2+]i during NO exposure was preceded by a decrease in mitochondrial membrane potential that was partly reversible during washout. Repeated applications of NMDA induced irreversible [Ca2+]i responses in a subpopulation of striatal cells that were greatly reduced by the NOS inhibitor N omega-nitro-l-arginine. Calcium transients were prolonged by conjoint application of NMDA and NO. We conclude that NMDA-evoked [Ca2+]i transients are modulated by endogenous NO production, which leads to release of calcium from the mitochondrial pool. An NO-activated mitochondrial permeability transition pore may lead to cell death after overstimulation of NMDA receptors.

Animals↗