Activation of Endogenous Protein Kinase C by Halothane in Synaptosomes

Hemmings, Hugh C.; Adamo, Anna I. B.

doi:10.1097/00000542-199603000-00021

Cited by 34 publications

(17 citation statements)

References 57 publications

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“…Regarding the G qcoupled receptors, we recently reported that halothane inhibits functions of 5-HT 2A R and M 1 R via a PKCdependent mechanism [2,4]. In vitro studies have also demonstrated that halothane affects PKC activity in the brain [26][27][28]. In our experimental system, M 2 R activation could activate PKC through a G qi5 -mediated PLC pathway.…”

mentioning

confidence: 59%

Analysis of the Effects of Halothane on Gi-Coupled Muscarinic M2 Receptor Signaling in Xenopus Oocytes Using a Chimeric Gα Protein

Minami

Uezono²,

Shiraishi³

et al. 2004

Pharmacology

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Metabotropic G protein-coupled receptors have recently been recognized as targets for anesthetics and analgesics. In particular, G_q-coupled receptors such as muscarinic M₁ receptors (M₁R) and 5-hydroxytryptamine (5-HT) type 2A receptors have been reported to be targets for anesthetics. Much less is known, however, about the effects of anesthetics on G_i-coupled receptors. Here we report a method to analyze functions of G_i-coupled receptors in Xenopus oocytes expressing a chimeric Gα protein. A chimeric Gα_q protein Gα_qi5, which contains carboxy-terminus five amino acids of Gα_i, enables G_i-coupled receptors to couple to Gq-coupled receptor-mediated downstream pathways such as activation of phospholipase C. We determined acetylcholine (ACh)-induced Ca²⁺-activated Cl^– currents in Xenopus oocytes coexpressing G_i-coupled muscarinic M₂receptors (M₂R) with the chimeric Gα_qi5. Although ACh did not induce any currents in oocytes expressing M₂R alone, it caused robust Cl^– currents in oocytes coexpressing M₂R with Gα_qi5. The EC₅₀ of the ACh-induced Cl^– current mediated through Gα_qi5 was 0.2 µmol/l, which was 2.2 times higher than that of the ACh-induced G protein-activated inwardly rectifying K⁺ currents activated by Gβγ subunits liberated from endogenously expressed Gα_i in Xenopus oocytes. Other G_i-coupled somatostatin type 2, 5-HT_1A and δ-opioid receptors, when coexpressed with Gα_qi5 in oocytes, also caused robust Ca²⁺-activated Cl^– currents. In oocytes coexpressing M₂R and Gα_qi5, a volatile anesthetic halothane inhibited M₂R-induced Cl^– currents in a concentration-dependent manner with the IC₅₀ of 1.1 mmol/l, suggesting that halothane inhibits M₂R-induced cellular responses at clinically relevant concentrations. Treatment with the protein kinase C inhibitor GF109203X produced a 3.5-fold enhancement of the initial Cl^– currents induced by 1 µmol/l ACh in oocytes expressing M₂R and G_qi5. The rate of halothane-induced inhibition of Cl^– currents elicited by ACh, however, was not changed in such oocytes pretreated with GF109203X. These findings suggest that halothane inhibits the M₂R-induced signaling by acting at sites other than PKC activity. Collectively these findings suggest that the use of oocyte expressing Gα_qi5 would be helpful to examine the effects of anesthetics or analgesics on the function of G_i-coupled receptors in the Xenopus oocyte expression system.

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mentioning

confidence: 59%

Analysis of the Effects of Halothane on Gi-Coupled Muscarinic M2 Receptor Signaling in Xenopus Oocytes Using a Chimeric Gα Protein

Minami

Uezono²,

Shiraishi³

et al. 2004

Pharmacology

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“…Halothane and propofol stimulate the activation of rat partial purified PKC in the presence of physiologically relevant bilayer vesicles in vitro [4]. Halothane also stimulates PKC activity in rat cerebrocortical synaptosomes [6]. Although these reports suggest that anesthetics may function in the brain through PKC signal transduction, little is known about the detailed effects of anesthetics on PKC signaling in neuronal cells.…”

Section: Introductionmentioning

confidence: 99%

Sevoflurane Stimulates MAP Kinase Signal transduction through the Activation of PKC .ALPHA. and .BETA.II in Fetal Rat Cerebral Cortex Cultured Neuron

Hasegawa

Takekoshi

Nagata

et al. 2006

Acta Histochem. Cytochem

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Protein kinase C (PKC) is a key enzyme that participates in various neuronal functions. PKC has also been identified as a target molecule for general anesthetic actions. Raf, mitogen-activated protein kinase (MEK) and extracellular signal-regulated kinase (ERK1/2) have been thought to be target effectors of PKC. In the present study, we attempted to evaluate the effect of sevoflurane on PKC/MAPK cascade signaling in cultured fetal rat cerebral cortex neurons, prepared from embryonic day 18 fetuses. The effects of sevoflurane on the translocation of 7 PKC isoforms (α, βI, βII, γ, δ, ɛ and ζ) were observed by immunoblotting using isoform-selective antibodies to PKCs. The treatment of neurons with sevoflurane induced the translocation of PKC α and PKC βII species from the cytosol to the membrane fraction, which indicated the activation of these PKC isoforms. In contrast, there was no clear change in the distribution of other PKC isoforms. We next examined whether the specific activation of PKC α and βII by sevoflurane could stimulate the MAP kinase signaling pathway in cultured neurons. Raf phosphorylation was increased by the administration of 0.25 mM sevoflurane. The phosphorylation of Raf proteins reached a maximum at 5–10 min. Subsequently, the phosphorylation of MEK proteins was increased at 10–15 min after sevoflurane treatments. That of ERK proteins was induced at 15–60 min. Moreover, the phosphorylation of ERK induced by sevoflurane was significantly decreased by the treatment of PKC inhibitor (staurosporine) and MEK inhibitor (PD98059). On the other hand, the contents of total Raf, MEK and ERK proteins were relatively constant at all times examined. To examine the localization of phosphorylated-ERK protein, immunohistochemical staining of sevoflurane-treated cultured neurons was performed. The phosphorylated-ERK proteins were markedly accumulated in both the cytosol of the cell body and the neurites in the neuronal cells with time after 0.25 mM sevoflurane-treatment. These results demonstrated that sevoflurane induced the phosphorylation of the MAP kinase cascade through the activation of the PKC α and PKC βII species.

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“…Halothane increases both purified (Hemmings and Adamo, 1994) and endogenous (Hemmings and Adamo, 1996) brain protein kinase C (PKC) activity. Phosphorylation of Na + channels by PKC and PKA reduces Na + channel activity by altering channel kinetics, for example by slowing inactivation, and is therefore an important component of neuromodulation (Cantrell and Catterall, 2001).…”

Section: Mechanisms Of General Anesthetic Effects On the Central Nervmentioning

confidence: 99%

Sodium Channels as Targets for Volatile Anesthetics

Herold¹,

Hemmings²

2012

Front. Pharmacol.

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The molecular mechanisms of modern inhaled anesthetics are still poorly understood although they are widely used in clinical settings. Considerable evidence supports effects on membrane proteins including ligand- and voltage-gated ion channels of excitable cells. Na+ channels are crucial to action potential initiation and propagation, and represent potential targets for volatile anesthetic effects on central nervous system depression. Inhibition of presynaptic Na+ channels leads to reduced neurotransmitter release at the synapse and could therefore contribute to the mechanisms by which volatile anesthetics produce their characteristic end points: amnesia, unconsciousness, and immobility. Early studies on crayfish and squid giant axon showed inhibition of Na+ currents by volatile anesthetics at high concentrations. Subsequent studies using native neuronal preparations and heterologous expression systems with various mammalian Na+ channel isoforms implicated inhibition of presynaptic Na+ channels in anesthetic actions at clinical concentrations. Volatile anesthetics reduce peak Na+ current (INa) and shift the voltage of half-maximal steady-state inactivation (h∞) toward more negative potentials, thus stabilizing the fast-inactivated state. Furthermore recovery from fast-inactivation is slowed, together with enhanced use-dependent block during pulse train protocols. These effects can depress presynaptic excitability, depolarization and Ca2+ entry, and ultimately reduce transmitter release. This reduction in transmitter release is more potent for glutamatergic compared to GABAergic terminals. Involvement of Na+ channel inhibition in mediating the immobility caused by volatile anesthetics has been demonstrated in animal studies, in which intrathecal infusion of the Na+ channel blocker tetrodotoxin increases volatile anesthetic potency, whereas infusion of the Na+ channels agonist veratridine reduces anesthetic potency. These studies indicate that inhibition of presynaptic Na+ channels by volatile anesthetics is involved in mediating some of their effects.

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Activation of Endogenous Protein Kinase C by Halothane in Synaptosomes

Cited by 34 publications

References 57 publications

Analysis of the Effects of Halothane on G<sub>i</sub>-Coupled Muscarinic M<sub>2</sub> Receptor Signaling in <i>Xenopus </i>Oocytes Using a Chimeric Gα Protein

Analysis of the Effects of Halothane on G<sub>i</sub>-Coupled Muscarinic M<sub>2</sub> Receptor Signaling in <i>Xenopus </i>Oocytes Using a Chimeric Gα Protein

Sevoflurane Stimulates MAP Kinase Signal transduction through the Activation of PKC .ALPHA. and .BETA.II in Fetal Rat Cerebral Cortex Cultured Neuron

Sodium Channels as Targets for Volatile Anesthetics

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