GABAA receptor function is regulated by phosphorylation in acutely dissociated guinea‐pig hippocampal neurones.

Chen, Qiang Xiao; Stelzer, Armin; Kay, Alan R.; Wong, Robert K. S.

doi:10.1113/jphysiol.1990.sp017908

Cited by 291 publications

(182 citation statements)

References 35 publications

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“…Along these lines, several studies have shown that GABA A receptor-mediated activity is regulated by phosphorylation. GABA A currents were decreased by Ͼ90% if cells were ATP-depleted (Chen et al, 1990), suggesting that a basal level of phosphorylation was required for maintenance of optimal GABA A receptor channel function. In addition, GABA A receptor conductance has been shown to be directly affected by PKA-mediated phosphorylation (Moss et al, 1992).…”

Section: Postsynaptic Effectsmentioning

confidence: 99%

GABA Activity Mediating Cytosolic Ca²⁺Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

Obrietan

Pol

1997

J. Neurosci.

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In the majority of developing neurons, GABA can exert depolarizing actions, thereby raising neuronal Ca 2ϩ . Ca 2ϩ elevations can have broad consequences during development, inducing gene expression, altering neurite outgrowth and growth cone turning, activating enzyme pathways, and influencing neuronal survival. We used fura-2 and fluo-3 Ca 2ϩ digital imaging to assess the effects of inhibiting or activating the cAMP signal transduction pathway on GABA activity mediating Ca 2ϩ rises during the early stages of in vitro hypothalamic neural development. Our experiments stemmed from the finding that stimulation of transmitter receptors shown to either activate or inhibit adenylyl cyclase activity caused a rapid decrease in Ca 2ϩ rises mediated by synaptically released GABA. Both the adenylyl cyclase activator forskolin and the inhibitor SQ-22,536 reduced the Ca 2ϩ rise elicited by the synaptic release of GABA. Bath application of the membrane-permeable cAMP analogs 8-bromo-cAMP (8-Br-cAMP) or 8-(4-chlorophenylthio)-cAMP (0.2-5 mM) produced a rapid, reversible, dose-dependent inhibition of Ca 2ϩ rises triggered by synaptic GABA release. Potentiation of GABAergic activity mediating Ca 2ϩ rises was observed in some neurons at relatively low concentrations of the membrane-permeable cAMP analogs (20 -50 M). In the presence of tetrodotoxin (TTX), postsynaptic Ca 2ϩ rises triggered by the bath application of GABA were only moderately depressed (13%) by 8-Br-cAMP (1 mM), suggesting that the inhibitory effects of 8-Br-cAMP were largely the result of a presynaptic mechanism.The protein kinase A (PKA) inhibitors H89 and Rp-3Ј,5Ј-cyclic monophosphothioate triethylamine also caused a large reduction (Ͼ70%) in Ca 2ϩ rises triggered by synaptic GABA release. Unlike the short-term depression elicited by activation of the cAMP signal transduction pathway, Ca 2ϩ depression elicited by PKA inhibition persisted for an extended period (Ͼ30 min) after PKA inhibitor washout. Postsynaptic depression of GABA-evoked Ca 2ϩ rises triggered by H89 (in the presence of TTX) recovered rapidly, suggesting that the extended depression observed during synaptic GABA release was largely through a presynaptic mechanism. Long-term Ca 2ϩ modulation by cAMP-regulating hypothalamic peptides may be mediated through a parallel mechanism.Together, these results suggest that GABAergic activity mediating Ca 2ϩ rises is dependent on ongoing PKA activity that is maintained within a narrow zone for GABA to elicit a maximal Ca 2ϩ elevation. Thus, neuromodulator-mediated changes in the cAMP-dependent signal transduction pathway (activation or inhibition) could lead to a substantial decrease in GABAmediated Ca 2ϩ rises during early development.

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Section: Postsynaptic Effectsmentioning

confidence: 99%

GABA Activity Mediating Cytosolic Ca²⁺Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

Obrietan

Pol

1997

J. Neurosci.

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“…Recordings were made with pipettes (resistance 2-5 M⍀) filled with intracellular solution that contained (in mM): CsCl 140, HEPES 10, 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid 10, pH 7.25; osmolarity, 300 -315 mOsm. Na 2 -ATP (4 mM) and MgCl 2 (2 mM) were included in the intracellular recording solution to support the process of protein phosphorylation, thereby preventing current rundown during a prolonged period of whole-cell recording (Chen et al, 1990). Currents were recorded under standard whole-cell voltage-clamp configuration using an Axopatch 1D amplifier (Axon Instruments, Foster City, CA).…”

Section: Methodsmentioning

confidence: 99%

Modulation of GABA_AReceptor Function by Tyrosine Phosphorylation of β Subunits

et al. 1997

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Protein tyrosine phosphorylation is a key event in diverse intracellular signaling pathways and has been implicated in modification of neuronal functioning. We investigated the role of tyrosine phosphorylation in regulating type A GABA (GABA A ) receptors in cultured CNS neurons. Extracellular application of genistein (50 M), a membrane-permeable inhibitor of protein tyrosine kinases (PTKs), produced a reversible reduction in the amplitude of GABA A receptor-mediated whole-cell currents, and this effect was not reproduced by daidzein (50 M), an inactive analog of genistein. In contrast, intracellular application of the PTK pp60 c-src (30 U/ml) resulted in a progressive increase in current amplitude, and this potentiation was prevented by pretreatment of the neurons with genistein. Immunoprecipitation and immunoblotting of cultured neuronal homogenates indicated that the ␤2/␤3 subunit(s) of the GABA A receptor are tyrosine phosphorylated in situ. Moreover, genistein (50 M) was found to be capable of decreasing GABA A currents in human embryonic kidney 293 cells transiently expressing functional GABA A receptors containing the ␤2 subunit. Thus, the present work provides the first evidence that native GABA A receptors are phosphorylated and modulated in situ by endogenous PTKs in cultured CNS neurons and that phosphorylation of the ␤ subunits may be sufficient to support such a modulation. Given the prominent role of GABA A receptors in mediating many brain functions and dysfunctions, modulation of these receptors by PTKs may be important in a wide range of physiological and pathological processes in the CNS.

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“…This rundown involves a phosphorylationdephosphorylation process (Stelzer et al, 1988;Chen et al, 1990;Gyenes et al, 1994;Kaneda et al, 1995). Using whole-cell recordings, the currents elicited by GABA (100 M) were measured in neurons acutely dissociated from rat cortex (Fig.…”

Section: Function Of Gapdh-dependent Phosphorylationmentioning

confidence: 99%

“…In acutely dissociated neurons or in slices, maintenance of GABA A R function requires favorable conditions for phosphorylation, by an unknown endogenous kinase requiring only the addition of ATP in the intracellular milieu (Stelzer et al, 1988;Chen et al, 1990;Krupp and Feltz, 1993;Gyenes et al, 1994;Kaneda et al, 1995). Using purified GABA A R from bovine cerebral cortex, we and others have previously shown that this receptor-associated kinase activity is ␣1 specific (Sweetnam et al, 1988;Bureau and Laschet, 1995).…”

Section: Introductionmentioning

confidence: 99%

Glyceraldehyde-3-Phosphate Dehydrogenase Is a GABA_AReceptor Kinase Linking Glycolysis to Neuronal Inhibition

Khallou-Laschet¹,

Minier²,

Kurcewicz³

et al. 2004

J. Neurosci.

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Protein phosphorylation is crucial for regulating synaptic transmission. We describe a novel mechanism for the phosphorylation of the GABA A receptor, which mediates fast inhibition in the brain. A protein copurified and coimmunoprecipitated with the phosphorylated receptor ␣1 subunit; this receptor-associated protein was identified by purification and microsequencing as the key glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Molecular constructs demonstrated that GAPDH directly phosphorylates the long intracellular loop of GABA A receptor ␣1 subunit at identified serine and threonine residues. GAPDH and the ␣1 subunit were found to be colocalized at the neuronal plasma membrane. In keeping with the GAPDH/GABA A receptor molecular association, glycolytic ATP produced locally at plasma membranes was consumed for this ␣1 subunit phosphorylation, possibly within a single macrocomplex. The membrane-attached GAPDH is thus a dual-purpose enzyme, a glycolytic dehydrogenase, and a receptor-associated kinase. In acutely dissociated cortical neurons, the rundown of the GABA A responses was essentially attributable to a Mg 2ϩ -dependent phosphatase activity, which was sensitive to vanadate but insensitive to okadaic acid or fluoride. Rundown was significantly reduced by the addition of GAPDH or its reduced cofactor NADH and nearly abolished by the addition of its substrate glyceraldehyde-3-phosphate (G3P). The prevention of rundown by G3P was abolished by iodoacetamide, an inhibitor of the dehydrogenase activity of GAPDH, indicating that the GABA A responses are maintained by a glycolysis-dependent phosphorylation. Our results provide a molecular mechanism for the direct involvement of glycolysis in neurotransmission.

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GABAA receptor function is regulated by phosphorylation in acutely dissociated guinea‐pig hippocampal neurones.

Cited by 291 publications

References 35 publications

GABA Activity Mediating Cytosolic Ca²⁺Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

GABA Activity Mediating Cytosolic Ca²⁺Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

Modulation of GABA_AReceptor Function by Tyrosine Phosphorylation of β Subunits

Glyceraldehyde-3-Phosphate Dehydrogenase Is a GABA_AReceptor Kinase Linking Glycolysis to Neuronal Inhibition

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GABAA receptor function is regulated by phosphorylation in acutely dissociated guinea‐pig hippocampal neurones.

Cited by 291 publications

References 35 publications

GABA Activity Mediating Cytosolic Ca2+Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

GABA Activity Mediating Cytosolic Ca2+Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

Modulation of GABAAReceptor Function by Tyrosine Phosphorylation of β Subunits

Glyceraldehyde-3-Phosphate Dehydrogenase Is a GABAAReceptor Kinase Linking Glycolysis to Neuronal Inhibition

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GABA Activity Mediating Cytosolic Ca²⁺Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

GABA Activity Mediating Cytosolic Ca²⁺Rises in Developing Neurons Is Modulated by cAMP-Dependent Signal Transduction

Modulation of GABA_AReceptor Function by Tyrosine Phosphorylation of β Subunits

Glyceraldehyde-3-Phosphate Dehydrogenase Is a GABA_AReceptor Kinase Linking Glycolysis to Neuronal Inhibition