A Common Molecular Mechanism for Active Emergence of Consciousness from Anesthesia

As early as the 1840, scientists began to explore the mechanisms of anesthesia: how could a simple small molecule lead to general anesthesia with loss of consciousness? Since then, the molecular and neural mechanisms of anesthesia-induced loss of consciousness have been the focus of research. However, in the past two decades, the mechanism of recovery of consciousness from anesthesia has become a new focus of research. For the past long period of time, it has been recognized that recovery of consciousness from anesthesia was simply regarded as the opposite process of induction of anesthesia, and consciousness returned when the concentration of the anesthetic in the body was reduced to a certain concentration. Nowadays, the idea that the recovery of consciousness is the reverse process of anesthesia has been challenged and people have realized that the recovery of consciousness cannot be simply attributed to the elimination of anesthetics. The recovery of consciousness is an active and controllable process, which is independent of elimination of anesthetics and has different neural mechanisms.

Because of the reversibility and controllability, anesthesia-induced loss of consciousness is considered as an ideal model to explore the recovery of consciousness. In the past decade, several neural circuits related to the recovery of consciousness have been revealed, and activation of these circuits through pharmacological or genetic techniques can accelerate the recovery of consciousness. However, the core mechanisms have not been revealed. If the recovery of consciousness is not a purely passive process, what mechanisms initiate and facilitate the active recovery of consciousness?

Our study focused mainly on exploring the common molecular mechanisms that promote the recovery of consciousness from anesthesia. In this study, a variety of experimental techniques were used, including western blot, immunofluorescence, two-photon brain slice imaging, patch clamp technique, stereotaxic technique, and in vivo electrical activity recording to investigate the role of K⁺-Cl^– cotransporter 2 (KCC2) ubiquitin degradation in recovery of consciousness. The results showed that KCC2 was downregulated significantly in both the hypothalamus and the thalamus during deep anesthesia (defined as the minimum responsive state, MRS) induced by different anesthetics, propofol, pentobarbitone, ketamine, and isoflurane, and this downregulated was triggered when the level of consciousness was close to the minimum responsive state. Furthermore, seven nuclei with the most significant changes in KCC2 expression in the thalamus and hypothalamus were screened by immunofluorescence assay. The results showed that only decreasing KCC2 expression in the ventral posteromedial nucleus of the thalamus could accelerate the recovery of consciousness.

KCC2 is a key molecule in the regulation of inhibitory GABAergic transmission in the matured central nervous system. When KCC2 expression is reduced, it results in a shift from inhibitory to excitatory GABAergic transmission. The results of patch clamp experiments showed that GABA-evoked potentials (E_GABA) were significantly shifted toward the positive direction after brain slices were treated with different anesthetics. Thus, KCC2 reduction-dependent GABA shift may be an intrinsic driving force for recovery of consciousness.

The detailed molecular mechanisms underlying the reduced expression of KCC2 in the minimum responsive state were also explored. Pharmacological inhibition of the ubiquitin proteasomal degradation pathway prevents anesthesia-induced decrease in KCC2, prolongs the duration of anesthesia, and delays recovery of consciousness from anesthesia. Substrate ubiquitination is the first key process in the ubiquitin proteasome pathway, and E3 ubiquitin ligase is one of the key molecules for ubiquitination. If KCC2 can be degraded by ubiquitin, then an E3 ubiquitin ligase that specifically recognizes KCC2 must exist. Therefore, it was necessary to identify E3 ligases that could recognize KCC2. Through proteomics and co-immunoprecipitation, we identified Fbxl4 as a specific ligase of KCC2. When Fbxl4, a member of the F-box protein family, is specifically altered in the ventral posteromedial nucleus of the thalamus (VPM), KCC2 expression and recovery of consciousness can be affected accordingly. Fbxl4 binds to the substrate by recognizing a structure called a "degron" on the substrate. This recognition process is regulated in ways that include substrate phosphorylation. Our further findings revealed that recognition of KCC2 by Fbxl4 is regulated by of KCC2 Thr1007 phosphorylation. In addition, transport of ubiquitinated substrates to the proteasome is the second key process in the ubiquitin-proteasome pathway. Our results showed that ubiquitinated KCC2 formed a complex with VCP with the assistance of FAF1 and was then transported to the proteasome for degradation.

KCC2 Thr1007 phosphorylation is a prerequisite for KCC2 ubiquitin degradation. When KCC2 phosphorylation was inhibited, KCC2 ubiquitination was also inhibited. N-Ethylmaleimide (NEM) was confirmed to enhance the activity of KCC2 by inhibiting its phosphorylation at Thr1007. Therefore, we preliminarily explored the role of NEM in the recovery of consciousness. Our study found that NEM prevented ubiquitin degradation of KCC2 and prolonged the duration of anesthesia.

This study is the first to reveal the molecular mechanism of the recovery of consciousness after anesthesia, confirming that the recovery of consciousness is an active process. When the level of consciousness approaches the minimum responsive state, the brain initiates the ubiquitin proteasome system to degrade KCC2, thus promoting the recovery of consciousness. The molecular mechanism of KCC2 ubiquitin degradation was investigated in detail, including the identification of the specific E3 ubiquitin ligase Fbxl4 and the key protein VCP responsible for the transport of ubiquitylated KCC2 to the proteasome. This study may also provide new ideas for the treatment of disorders of consciousness.

[1] KOCH, C., What is consciousness? [J]. Nature, 2018. 557(7704): p. S8-S12.
[2] BROWN, R., H. LAU, AND J.E. LEDOUX, Understanding the higher-order approach to consciousness[J]. Trends Cogn Sci, 2019. 23(9): p. 754-768.
[3] TONONI, G., et al., Integrated information theory: from consciousness to its physical substrate[J]. Nat Rev Neurosci, 2016. 17(7): p. 450-61.
[4] ROURK, C., Application of the catecholaminergic neuron electron transport (CNET) physical substrate for consciousness and action selection to integrated information theory[J]. Entropy (Basel), 2022. 24(1).
[5] PASCUAL-LEONE, A. AND V. WALSH, Fast backprojections from the motion to the primary visual area necessary for visual awareness[J]. Science, 2001. 292(5516): p. 510-2.
[6] MASHOUR, G.A., et al., Conscious processing and the global neuronal workspace hypothesis[J]. Neuron, 2020. 105(5): p. 776-798.
[7] NORTHOFF, G. AND F. ZILIO, Temporo-spatial theory of consciousness (TTC) - bridging the gap of neuronal activity and phenomenal states[J]. Behav Brain Res, 2022. 424: p. 113788.
[8] BLACK, D., The global workspace theory, the phenomenal concept strategy, and the distribution of consciousness[J]. Conscious Cogn, 2020. 84: p. 102992.
[9] LAU, H. AND D. ROSENTHAL, Empirical support for higher-order theories of conscious awareness[J]. Trends Cogn Sci, 2011. 15(8): p. 365-73.
[10] OIZUMI, M., L. ALBANTAKIS, AND G. TONONI, from the phenomenology to the mechanisms of consciousness: integrated information theory 3.0[J]. PLoS Comput Biol, 2014. 10(5): p. e1003588.
[11] TONONI, G. AND C. KOCH, Consciousness: here, there and everywhere? [J]. Philos Trans R Soc Lond B Biol Sci, 2015. 370(1668).
[12] SETH, A.K. AND T. BAYNE, Theories of consciousness[J]. Nat Rev Neurosci, 2022. 23(7): p. 439-452.
[13] DENNETT, D.C., Facing up to the hard question of consciousness[J]. Philos Trans R Soc Lond B Biol Sci, 2018. 373(1755).
[14] LI, Y., et al., Hypothalamic circuits for predation and evasion[J]. Neuron, 2018. 97(4): p. 911-924 e5.
[15] LIU, M., et al., Make war not love: the neural substrate underlying a state-dependent switch in female social behavior[J]. Neuron, 2021.
[16] DORRIES, K.M., Olfactory "consciousness"? [J]. Science, 1997. 278(5343): p. 1550.
[17] ARZI, A., et al., Olfactory sniffing signals consciousness in unresponsive patients with brain injuries[J]. Nature, 2020. 581(7809): p. 428-433.
[18] MIYAMICHI, K., et al., Cortical representations of olfactory input by trans-synaptic tracing[J]. Nature, 2011. 472(7342): p. 191-6.
[19] MERRICK, C., et al., The olfactory system as the gateway to the neural correlates of consciousness[J]. Front Psychol, 2014. 4: p. 1011.
[20] HALASSA, M.M. AND S. KASTNER, Thalamic functions in distributed cognitive control[J]. Nat Neurosci, 2017. 20(12): p. 1669-1679.
[21] EGNER, T. AND J. HIRSCH, Cognitive control mechanisms resolve conflict through cortical amplification of task-relevant information[J]. Nat Neurosci, 2005. 8(12): p. 1784-90.
[22] COLE, M.W., et al., Activity flow over resting-state networks shapes cognitive task activations[J]. Nat Neurosci, 2016. 19(12): p. 1718-1726.
[23] WHITTINGTON, J.C.R., et al., How to build a cognitive map[J]. Nat Neurosci, 2022. 25(10): p. 1257-1272.
[24] ETKIN, A., T. EGNER, AND R. KALISCH, Emotional processing in anterior cingulate and medial prefrontal cortex[J]. Trends Cogn Sci, 2011. 15(2): p. 85-93.
[25] EUGENE, F., et al., Neural correlates of inhibitory deficits in depression[J]. Psychiatry Res, 2010. 181(1): p. 30-5.
[26] HELLER, A.S., et al., Reduced capacity to sustain positive emotion in major depression reflects diminished maintenance of fronto-striatal brain activation[J]. Proc Natl Acad Sci U S A, 2009. 106(52): p. 22445-50.
[27] RUSSO, S.J. AND E.J. NESTLER, The brain reward circuitry in mood disorders[J]. Nat Rev Neurosci, 2013. 14(9): p. 609-25.
[28] SIKH, S.S. AND P.N. DHULIA, Recovery from general anesthesia: a simple and comprehensive test for assessment[J]. Anesth Analg, 1979. 58(4): p. 324-6.
[29] FRIEDMAN, E.B., et al., A conserved behavioral state barrier impedes transitions between anesthetic-induced unconsciousness and wakefulness: evidence for neural inertia[J]. PLoS One, 2010. 5(7): p. e11903.
[30] FRANKS, N.P. AND W.R. LIEB, Molecular and cellular mechanisms of general anaesthesia[J]. Nature, 1994. 367(6464): p. 607-14.
[31] FRANKS, N.P., General anaesthesia: from molecular targets to neuronal pathways of sleep and arousal[J]. Nat Rev Neurosci, 2008. 9(5): p. 370-86.
[32] KROM, A.J., et al., Anesthesia-induced loss of consciousness disrupts auditory responses beyond primary cortex[J]. Proc Natl Acad Sci U S A, 2020. 117(21): p. 11770-11780.
[33] HUDSON, A.E., et al., Recovery of consciousness is mediated by a network of discrete metastable activity states[J]. Proc Natl Acad Sci U S A, 2014. 111(25): p. 9283-8.
[34] ALKIRE, M.T., et al., Thalamic microinjection of nicotine reverses sevoflurane-induced loss of righting reflex in the rat[J]. Anesthesiology, 2007. 107(2): p. 264-72.
[35] HUDETZ, A.G., J.D. WOOD, AND J.P. KAMPINE, Cholinergic reversal of isoflurane anesthesia in rats as measured by cross-approximate entropy of the electroencephalogram[J]. Anesthesiology, 2003. 99(5): p. 1125-31.
[36] PAL, D., et al., Differential role of prefrontal and parietal cortices in controlling level of consciousness[J]. Curr Biol, 2018. 28(13): p. 2145-2152 e5.
[37] LUO, T. AND L.S. LEUNG, Basal forebrain histaminergic transmission modulates electroencephalographic activity and emergence from isoflurane anesthesia[J]. Anesthesiology, 2009. 111(4): p. 725-33.
[38] HNASKO, T.S., B.N. SOTAK, AND R.D. PALMITER, Morphine reward in dopamine-deficient mice[J]. Nature, 2005. 438(7069): p. 854-7.
[39] LI, N. AND A. JASANOFF, Local and global consequences of reward-evoked striatal dopamine release[J]. Nature, 2020. 580(7802): p. 239-244.
[40] ROBBINS, T.W., Dopamine and cognition[J]. Curr Opin Neurol, 2003. 16 Suppl 2: p. S1-2.
[41] TORRISI, S.A., et al., Dopamine D3 receptor, cognition and cognitive dysfunctions in neuropsychiatric disorders: from the bench to the bedside[J]. Curr Top Behav Neurosci, 2023. 60: p. 133-156.
[42] TAYLOR, N.E., et al., Activation of D1 dopamine receptors induces emergence from isoflurane general anesthesia[J]. Anesthesiology, 2013. 118(1): p. 30-9.
[43] SOLT, K., et al., Electrical stimulation of the ventral tegmental area induces reanimation from general anesthesia[J]. Anesthesiology, 2014. 121(2): p. 311-9.
[44] YU, X., et al., GABA and glutamate neurons in the VTA regulate sleep and wakefulness[J]. Nat Neurosci, 2019. 22(1): p. 106-119.
[45] TAYLOR, N.E., et al., Optogenetic activation of dopamine neurons in the ventral tegmental area induces reanimation from general anesthesia[J]. Proc Natl Acad Sci U S A, 2016. 113(45): p. 12826-12831.
[46] VAZEY, E.M. AND G. ASTON-JONES, Designer receptor manipulations reveal a role of the locus coeruleus noradrenergic system in isoflurane general anesthesia[J]. Proc Natl Acad Sci U S A, 2014. 111(10): p. 3859-64.
[47] PILLAY, S., et al., Norepinephrine infusion into nucleus basalis elicits microarousal in desflurane-anesthetized rats[J]. Anesthesiology, 2011. 115(4): p. 733-42.
[48] PANULA, P., H.Y. YANG, AND E. COSTA, Histamine-containing neurons in the rat hypothalamus[J]. Proc Natl Acad Sci U S A, 1984. 81(8): p. 2572-6.
[1] KOCH, C., What is consciousness?[J]. Nature, 2018. 557(7704): p. S8-S12.
[2] BROWN, R., H. LAU, AND J.E. LEDOUX, Understanding the higher-order approach to consciousness[J]. Trends Cogn Sci, 2019. 23(9): p. 754-768.
[3] TONONI, G., et al., Integrated information theory: from consciousness to its physical substrate[J]. Nat Rev Neurosci, 2016. 17(7): p. 450-61.
[4] ROURK, C., Application of the catecholaminergic neuron electron transport (CNET) physical substrate for consciousness and action selection to integrated information theory[J]. Entropy (Basel), 2022. 24(1).
[5] PASCUAL-LEONE, A. AND V. WALSH, Fast backprojections from the motion to the primary visual area necessary for visual awareness[J]. Science, 2001. 292(5516): p. 510-2.
[6] MASHOUR, G.A., et al., Conscious processing and the global neuronal workspace hypothesis[J]. Neuron, 2020. 105(5): p. 776-798.
[7] NORTHOFF, G. AND F. ZILIO, Temporo-spatial theory of consciousness (TTC) - bridging the gap of neuronal activity and phenomenal states[J]. Behav Brain Res, 2022. 424: p. 113788.
[8] BLACK, D., The global workspace theory, the phenomenal concept strategy, and the distribution of consciousness[J]. Conscious Cogn, 2020. 84: p. 102992.
[9] LAU, H. AND D. ROSENTHAL, Empirical support for higher-order theories of conscious awareness[J]. Trends Cogn Sci, 2011. 15(8): p. 365-73.
[10] OIZUMI, M., L. ALBANTAKIS, AND G. TONONI, from the phenomenology to the mechanisms of consciousness: integrated information theory 3.0[J]. PLoS Comput Biol, 2014. 10(5): p. e1003588.
[11] TONONI, G. AND C. KOCH, Consciousness: here, there and everywhere?[J]. Philos Trans R Soc Lond B Biol Sci, 2015. 370(1668).
[12] SETH, A.K. AND T. BAYNE, Theories of consciousness[J]. Nat Rev Neurosci, 2022. 23(7): p. 439-452.
[13] DENNETT, D.C., Facing up to the hard question of consciousness[J]. Philos Trans R Soc Lond B Biol Sci, 2018. 373(1755).
[14] LI, Y., et al., Hypothalamic circuits for predation and evasion[J]. Neuron, 2018. 97(4): p. 911-924 e5.
[15] LIU, M., et al., Make war not love: the neural substrate underlying a state-dependent switch in female social behavior[J]. Neuron, 2021.
[16] DORRIES, K.M., Olfactory "consciousness"?[J]. Science, 1997. 278(5343): p. 1550.
[17] ARZI, A., et al., Olfactory sniffing signals consciousness in unresponsive patients with brain injuries[J]. Nature, 2020. 581(7809): p. 428-433.
[18] MIYAMICHI, K., et al., Cortical representations of olfactory input by trans-synaptic tracing[J]. Nature, 2011. 472(7342): p. 191-6.
[19] MERRICK, C., et al., The olfactory system as the gateway to the neural correlates of consciousness[J]. Front Psychol, 2014. 4: p. 1011.
[20] HALASSA, M.M. AND S. KASTNER, Thalamic functions in distributed cognitive control[J]. Nat Neurosci, 2017. 20(12): p. 1669-1679.
[21] EGNER, T. AND J. HIRSCH, Cognitive control mechanisms resolve conflict through cortical amplification of task-relevant information[J]. Nat Neurosci, 2005. 8(12): p. 1784-90.
[22] COLE, M.W., et al., Activity flow over resting-state networks shapes cognitive task activations[J]. Nat Neurosci, 2016. 19(12): p. 1718-1726.
[23] WHITTINGTON, J.C.R., et al., How to build a cognitive map[J]. Nat Neurosci, 2022. 25(10): p. 1257-1272.
[24] ETKIN, A., T. EGNER, AND R. KALISCH, Emotional processing in anterior cingulate and medial prefrontal cortex[J]. Trends Cogn Sci, 2011. 15(2): p. 85-93.
[25] EUGENE, F., et al., Neural correlates of inhibitory deficits in depression[J]. Psychiatry Res, 2010. 181(1): p. 30-5.
[26] HELLER, A.S., et al., Reduced capacity to sustain positive emotion in major depression reflects diminished maintenance of fronto-striatal brain activation[J]. Proc Natl Acad Sci U S A, 2009. 106(52): p. 22445-50.
[27] RUSSO, S.J. AND E.J. NESTLER, The brain reward circuitry in mood disorders[J]. Nat Rev Neurosci, 2013. 14(9): p. 609-25.
[28] SIKH, S.S. AND P.N. DHULIA, Recovery from general anesthesia: a simple and comprehensive test for assessment[J]. Anesth Analg, 1979. 58(4): p. 324-6.
[29] FRIEDMAN, E.B., et al., A conserved behavioral state barrier impedes transitions between anesthetic-induced unconsciousness and wakefulness: evidence for neural inertia[J]. PLoS One, 2010. 5(7): p. e11903.
[30] FRANKS, N.P. AND W.R. LIEB, Molecular and cellular mechanisms of general anaesthesia[J]. Nature, 1994. 367(6464): p. 607-14.
[31] FRANKS, N.P., General anaesthesia: from molecular targets to neuronal pathways of sleep and arousal[J]. Nat Rev Neurosci, 2008. 9(5): p. 370-86.
[32] KROM, A.J., et al., Anesthesia-induced loss of consciousness disrupts auditory responses beyond primary cortex[J]. Proc Natl Acad Sci U S A, 2020. 117(21): p. 11770-11780.
[33] HUDSON, A.E., et al., Recovery of consciousness is mediated by a network of discrete metastable activity states[J]. Proc Natl Acad Sci U S A, 2014. 111(25): p. 9283-8.
[34] ALKIRE, M.T., et al., Thalamic microinjection of nicotine reverses sevoflurane-induced loss of righting reflex in the rat[J]. Anesthesiology, 2007. 107(2): p. 264-72.
[35] HUDETZ, A.G., J.D. WOOD, AND J.P. KAMPINE, Cholinergic reversal of isoflurane anesthesia in rats as measured by cross-approximate entropy of the electroencephalogram[J]. Anesthesiology, 2003. 99(5): p. 1125-31.
[36] PAL, D., et al., Differential role of prefrontal and parietal cortices in controlling level of consciousness[J]. Curr Biol, 2018. 28(13): p. 2145-2152 e5.
[37] LUO, T. AND L.S. LEUNG, Basal forebrain histaminergic transmission modulates electroencephalographic activity and emergence from isoflurane anesthesia[J]. Anesthesiology, 2009. 111(4): p. 725-33.
[38] HNASKO, T.S., B.N. SOTAK, AND R.D. PALMITER, Morphine reward in dopamine-deficient mice[J]. Nature, 2005. 438(7069): p. 854-7.
[39] LI, N. AND A. JASANOFF, Local and global consequences of reward-evoked striatal dopamine release[J]. Nature, 2020. 580(7802): p. 239-244.
[40] ROBBINS, T.W., Dopamine and cognition[J]. Curr Opin Neurol, 2003. 16 Suppl 2: p. S1-2.
[41] TORRISI, S.A., et al., Dopamine D3 receptor, cognition and cognitive dysfunctions in neuropsychiatric disorders: from the bench to the bedside[J]. Curr Top Behav Neurosci, 2023. 60: p. 133-156.
[42] TAYLOR, N.E., et al., Activation of D1 dopamine receptors induces emergence from isoflurane general anesthesia[J]. Anesthesiology, 2013. 118(1): p. 30-9.
[43] SOLT, K., et al., Electrical stimulation of the ventral tegmental area induces reanimation from general anesthesia[J]. Anesthesiology, 2014. 121(2): p. 311-9.
[44] YU, X., et al., GABA and glutamate neurons in the VTA regulate sleep and wakefulness[J]. Nat Neurosci, 2019. 22(1): p. 106-119.
[45] TAYLOR, N.E., et al., Optogenetic activation of dopamine neurons in the ventral tegmental area induces reanimation from general anesthesia[J]. Proc Natl Acad Sci U S A, 2016. 113(45): p. 12826-12831.
[46] VAZEY, E.M. AND G. ASTON-JONES, Designer receptor manipulations reveal a role of the locus coeruleus noradrenergic system in isoflurane general anesthesia[J]. Proc Natl Acad Sci U S A, 2014. 111(10): p. 3859-64.
[47] PILLAY, S., et al., Norepinephrine infusion into nucleus basalis elicits microarousal in desflurane-anesthetized rats[J]. Anesthesiology, 2011. 115(4): p. 733-42.
[48] PANULA, P., H.Y. YANG, AND E. COSTA, Histamine-containing neurons in the rat hypothalamus[J]. Proc Natl Acad Sci U S A, 1984. 81(8): p. 2572-6.
[49] SHIRASAKA, T., et al., Effects of orexin-A on propofol anesthesia in rats[J]. J Anesth, 2011. 25(1): p. 65-71.
[50] TOSE, R., ET AL., Orexin A decreases ketamine-induced anesthesia time in the rat: the relevance to brain noradrenergic neuronal activity[J]. Anesth Analg, 2009. 108(2): p. 491-5.
[51] KELZ, M.B., et al., An essential role for orexins in emergence from general anesthesia[J]. Proc Natl Acad Sci U S A, 2008. 105(4): p. 1309-14.
[52] KAILA, K., et al., Cation-chloride cotransporters in neuronal development, plasticity and disease[J]. Nature Reviews Neuroscience, 2014. 15(10): p. 637-654.
[53] BARTHO, P., et al., Differential distribution of the KCl cotransporter KCC2 in thalamic relay and reticular nuclei[J]. Eur J Neurosci, 2004. 20(4): p. 965-75.
[54] RIVERA, C., et al., The K+/Cl- co-transporter KCC2 renders GABA hyperpolarizing during neuronal maturation[J]. Nature, 1999. 397(6716): p. 251-5.
[55] DELPIRE, E., Cation-chloride cotransporters in neuronal communication[J]. News Physiol Sci, 2000. 15: p. 309-312.
[56] HU, J.J., et al., Bumetanide reduces the seizure susceptibility induced by pentylenetetrazol via inhibition of aberrant hippocampal neurogenesis in neonatal rats after hypoxia-ischemia[J]. Brain Res Bull, 2017. 130: p. 188-199.
[57] DZHALA, V.I., et al., NKCC1 transporter facilitates seizures in the developing brain[J]. Nat Med, 2005. 11(11): p. 1205-13.
[58] BOULENGUEZ, P., et al., Down-regulation of the potassium-chloride cotransporter KCC2 contributes to spasticity after spinal cord injury[J]. Nat Med, 2010. 16(3): p. 302-7.
[59] HASBARGEN, T., et al., Role of NKCC1 and KCC2 in the development of chronic neuropathic pain following spinal cord injury[J]. Ann N Y Acad Sci, 2010. 1198: p. 168-72.
[60] EFTEKHARI, S., et al., Bumetanide reduces seizure frequency in patients with temporal lobe epilepsy[J]. Epilepsia, 2013. 54(1): p. e9-12.
[61] HINZ, L., J. TORRELLA BARRUFET, AND V.M. HEINE, KCC2 expression levels are reduced in postmortem brain tissue of Rett syndrome patients[J]. Acta Neuropathol Commun, 2019. 7(1): p. 196.
[62] HUBERFELD, G., et al., Perturbed chloride homeostasis and GABAergic signaling in human temporal lobe epilepsy[J]. J Neurosci, 2007. 27(37): p. 9866-73.
[63] KAHLE, K.T., et al., Genetically encoded impairment of neuronal KCC2 cotransporter function in human idiopathic generalized epilepsy[J]. EMBO Rep, 2014. 15(7): p. 766-74.
[64] BUHL, E.H., T.S. OTIS, AND I. MODY, Zinc-induced collapse of augmented inhibition by GABA in a temporal lobe epilepsy model[J]. Science, 1996. 271(5247): p. 369-73.
[65] PATHAK, H.R., et al., Disrupted dentate granule cell chloride regulation enhances synaptic excitability during development of temporal lobe epilepsy[J]. J Neurosci, 2007. 27(51): p. 14012-22.
[66] VIITANEN, T., et al., The K+-Cl– cotransporter KCC2 promotes GABAergic excitation in the mature rat hippocampus[J]. J Physiol, 2010. 588(Pt 9): p. 1527-40.
[67] COHEN, I., et al., On the origin of interictal activity in human temporal lobe epilepsy in vitro[J]. Science, 2002. 298(5597): p. 1418-21.
[68] PUSKARJOV, M., et al., A variant of KCC2 from patients with febrile seizures impairs neuronal Cl- extrusion and dendritic spine formation[J]. EMBO Rep, 2014. 15(6): p. 723-9.
[69] FERRINI, F., et al., Morphine hyperalgesia gated through microglia-mediated disruption of neuronal Cl(-) homeostasis[J]. Nat Neurosci, 2013. 16(2): p. 183-92.
[70] LEE, H.H., et al., NMDA receptor activity downregulates KCC2 resulting in depolarizing GABAA receptor-mediated currents[J]. Nat Neurosci, 2011. 14(6): p. 736-43.
[71] RIVERA, C., et al., BDNF-induced TrkB activation down-regulates the K+-Cl- cotransporter KCC2 and impairs neuronal Cl- extrusion[J]. J Cell Biol, 2002. 159(5): p. 747-52.
[72] RIVERA, C., et al., Mechanism of activity-dependent downregulation of the neuron-specific K-Cl cotransporter KCC2[J]. J Neurosci, 2004. 24(19): p. 4683-91.
[73] GAGNON, M., et al., Chloride extrusion enhancers as novel therapeutics for neurological diseases[J]. Nat Med, 2013. 19(11): p. 1524-8.
[74] LI, H., et al., KCC2 interacts with the dendritic cytoskeleton to promote spine development[J]. Neuron, 2007. 56(6): p. 1019-33.
[75] FIUMELLI, H., et al., An ion transport-independent role for the cation-chloride cotransporter KCC2 in dendritic spinogenesis in vivo[J]. Cereb Cortex, 2013. 23(2): p. 378-88.
[76] HERING, H. AND M. SHENG, Dendritic spines: structure, dynamics and regulation[J]. Nat Rev Neurosci, 2001. 2(12): p. 880-8.
[77] HORN, Z., et al., Premature expression of KCC2 in embryonic mice perturbs neural development by an ion transport-independent mechanism[J]. Eur J Neurosci, 2010. 31(12): p. 2142-55.
[78] PUSKARJOV, M., et al., K-Cl Cotransporter 2-mediated Cl- extrusion determines developmental stage-dependent impact of propofol anesthesia on dendritic spines[J]. Anesthesiology, 2017. 126(5): p. 855-867.
[79] RUDOLPH, U. AND B. ANTKOWIAK, Molecular and neuronal substrates for general anaesthetics[J]. Nat Rev Neurosci, 2004. 5(9): p. 709-20.
[80] MILLER, K.W., Anaesthesia. Models of consciousness[J]. Nature, 1986. 323(6089): p. 584.
[81] SWATEK, K.N. AND D. KOMANDER, Ubiquitin modifications[J]. Cell Res, 2016. 26(4): p. 399-422.
[82] ROOS-MATTJUS, P. AND L. SISTONEN, The ubiquitin-proteasome pathway[J]. Ann Med, 2004. 36(4): p. 285-95.
[83] NANDI, D., et al., The ubiquitin-proteasome system[J]. J Biosci, 2006. 31(1): p. 137-55.
[84] LI, W., Et al., Genome-wide and functional annotation of human E3 ubiquitin ligases identifies MULAN, a mitochondrial E3 that regulates the organelle's dynamics and signaling[J]. PLoS One, 2008. 3(1): p. e1487.
[85] MORREALE, F.E. AND H. WALDEN, Types of Ubiquitin Ligases[J]. Cell, 2016. 165(1): p. 248-248 e1.
[86] KANTAMNENI, S., K.A. WILKINSON, AND J.M. HENLEY, Ubiquitin regulation of neuronal excitability[J]. Nat Neurosci, 2011. 14(2): p. 126-8.
[87] MABB, A.M. AND M.D. EHLERS, Ubiquitination in postsynaptic function and plasticity[J]. Annu Rev Cell Dev Biol, 2010. 26: p. 179-210.
[88] BERGOLD, P.J., et al., Protein synthesis during acquisition of long-term facilitation is needed for the persistent loss of regulatory subunits of the Aplysia cAMP-dependent protein kinase[J]. Proc Natl Acad Sci U S A, 1990. 87(10): p. 3788-91.
[89] CHAIN, D.G., et al., Mechanisms for generating the autonomous cAMP-dependent protein kinase required for long-term facilitation in Aplysia[J]. Neuron, 1999. 22(1): p. 147-56.
[90] JIANG, Y.H., et al., Mutation of the Angelman ubiquitin ligase in mice causes increased cytoplasmic p53 and deficits of contextual learning and long-term potentiation[J]. Neuron, 1998. 21(4): p. 799-811.
[91] LI, M., et al., The adaptor protein of the anaphase promoting complex Cdh1 is essential in maintaining replicative lifespan and in learning and memory[J]. Nat Cell Biol, 2008. 10(9): p. 1083-9.
[92] FU, A.K., et al., APC(Cdh1) mediates EphA4-dependent downregulation of AMPA receptors in homeostatic plasticity[J]. Nat Neurosci, 2011. 14(2): p. 181-9.
[93] MORIYOSHI, K., et al., Seven in absentia homolog 1A mediates ubiquitination and degradation of group 1 metabotropic glutamate receptors[J]. Proc Natl Acad Sci U S A, 2004. 101(23): p. 8614-9.
[94] ISHIKAWA, K., et al., Competitive interaction of seven in absentia homolog-1A and Ca2+/calmodulin with the cytoplasmic tail of group 1 metabotropic glutamate receptors[J]. Genes Cells, 1999. 4(7): p. 381-90.
[95] ALTIER, C., et al., The Cavbeta subunit prevents RFP2-mediated ubiquitination and proteasomal degradation of L-type channels[J]. Nat Neurosci, 2011. 14(2): p. 173-80.
[96] POPOVIC, D., D. VUCIC, AND I. DIKIC, Ubiquitination in disease pathogenesis and treatment[J]. Nat Med, 2014. 20(11): p. 1242-53.
[97] SPRATT, D.E., et al., A molecular explanation for the recessive nature of parkin-linked Parkinson's disease[J]. Nat Commun, 2013. 4: p. 1983.
[98] WANG, Y., et al., Synergy and antagonism of macroautophagy and chaperone-mediated autophagy in a cell model of pathological tau aggregation[J]. Autophagy, 2010. 6(1): p. 182-3.
[99] DESHAIES, R.J. AND C.A. JOAZEIRO, RING domain E3 ubiquitin ligases[J]. Annu Rev Biochem, 2009. 78: p. 399-434.
[100] CARDOZO, T. AND M. PAGANO, The SCF ubiquitin ligase: insights into a molecular machine[J]. Nat Rev Mol Cell Biol, 2004. 5(9): p. 739-51.
[101] MASON, B. AND H. LAMAN, The FBXL family of F-box proteins: variations on a theme[J]. Open Biol, 2020. 10(11): p. 200319.
[102] SKAAR, J.R., J.K. PAGAN, AND M. PAGANO, Mechanisms and function of substrate recruitment by F-box proteins[J]. Nat Rev Mol Cell Biol, 2013. 14(6): p. 369-81.
[103] CARRANO, A.C., et al., SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27[J]. Nat Cell Biol, 1999. 1(4): p. 193-9.
[104] SHEARD, L.B., et al., Jasmonate perception by inositol-phosphate-potentiated COI1-JAZ co-receptor[J]. Nature, 2010. 468(7322): p. 400-5.
[105] TAN, X., et al., Mechanism of auxin perception by the TIR1 ubiquitin ligase[J]. Nature, 2007. 446(7136): p. 640-5.
[106] ZENG, Z., et al., Structural basis of selective ubiquitination of TRF1 by SCFFbx4[J]. Dev Cell, 2010. 18(2): p. 214-25.
[107] LI, Y. AND B. HAO, Structural basis of dimerization-dependent ubiquitination by the SCF(Fbx4) ubiquitin ligase[J]. J Biol Chem, 2010. 285(18): p. 13896-906.
[108] YE, Y., H.H. MEYER, AND T.A. RAPOPORT, The AAA ATPase Cdc48/p97 and its partners transport proteins from the ER into the cytosol[J]. Nature, 2001. 414(6864): p. 652-6.
[109] RABINOVICH, E., et al., AAA-ATPase p97/Cdc48p, a cytosolic chaperone required for endoplasmic reticulum-associated protein degradation[J]. Mol Cell Biol, 2002. 22(2): p. 626-34.
[110] KLOPPSTECK, P., et al., Regulation of p97 in the ubiquitin-proteasome system by the UBX protein-family[J]. Biochim Biophys Acta, 2012. 1823(1): p. 125-9.
[111] ALEXANDRU, G., et al., UBXD7 binds multiple ubiquitin ligases and implicates p97 in HIF1alpha turnover[J]. Cell, 2008. 134(5): p. 804-16.
[112] ABILDGAARD, A.B., et al., Co-chaperones in targeting and delivery of misfolded proteins to the 26S proteasome[J]. Biomolecules, 2020. 10(8).
[113] KRIEGENBURG, F., L. ELLGAARD, AND R. HARTMANN-PETERSEN, Molecular chaperones in targeting misfolded proteins for ubiquitin-dependent degradation[J]. FEBS J, 2012. 279(4): p. 532-42.
[114] KANDASAMY, G. AND C. ANDREASSON, Hsp70-Hsp110 chaperones deliver ubiquitin-dependent and -independent substrates to the 26S proteasome for proteolysis in yeast[J]. J Cell Sci, 2018. 131(6).
[115] MANDAL, A.K., et al., Hsp110 chaperones control client fate determination in the hsp70-Hsp90 chaperone system[J]. Mol Biol Cell, 2010. 21(9): p. 1439-48.
[116] LUDERS, J., J. DEMAND, AND J. HOHFELD, The ubiquitin-related BAG-1 provides a link between the molecular chaperones Hsc70/Hsp70 and the proteasome[J]. J Biol Chem, 2000. 275(7): p. 4613-7.
[117] DEMAND, J., et al., Cooperation of a ubiquitin domain protein and an E3 ubiquitin ligase during chaperone/proteasome coupling[J]. Curr Biol, 2001. 11(20): p. 1569-77.
[118] BALLINGER, C.A., et al., Identification of CHIP, a novel tetratricopeptide repeat-containing protein that interacts with heat shock proteins and negatively regulates chaperone functions[J]. Mol Cell Biol, 1999. 19(6): p. 4535-45.
[119] KADOWAKI, H., et al., Pre-emptive quality control protects the ER from protein overload via the proximity of ERAD components and SRP[J]. Cell Rep, 2015. 13(5): p. 944-56.
[120] MINAMI, R., et al., BAG-6 is essential for selective elimination of defective proteasomal substrates[J]. J Cell Biol, 2010. 190(4): p. 637-50.
[121] MOCK, J.Y., et al., Bag6 complex contains a minimal tail-anchor-targeting module and a mock BAG domain[J]. Proc Natl Acad Sci U S A, 2015. 112(1): p. 106-11.
[122] KIKUKAWA, Y., et al., Unique proteasome subunit Xrpn10c is a specific receptor for the antiapoptotic ubiquitin-like protein Scythe[J]. FEBS J, 2005. 272(24): p. 6373-86.
[123] ZHANG, J., et al., Cyclin D-CDK4 kinase destabilizes PD-L1 via cullin 3-SPOP to control cancer immune surveillance[J]. Nature, 2018. 553(7686): p. 91-95.
[124] WEI, S., et al., Role of human Keap1 S53 and S293 residues in modulating the binding of Keap1 to Nrf2[J]. Biochimie, 2019. 158: p. 73-81.
[125] ZHANG, T., et al., G-protein-coupled receptors regulate autophagy by ZBTB16-mediated ubiquitination and proteasomal degradation of Atg14L[J]. Elife, 2015. 4: p. e06734.
[126] INUZUKA, H., et al., Phosphorylation by casein kinase I promotes the turnover of the Mdm2 oncoprotein via the SCF (beta-TRCP) ubiquitin ligase[J]. Cancer Cell, 2010. 18(2): p. 147-59.
[127] ABBAS, T., et al., CRL1-FBXO11 promotes Cdt2 ubiquitylation and degradation and regulates Pr-Set7/Set8-mediated cellular migration[J]. Mol Cell, 2013. 49(6): p. 1147-58.
[128] ROSSI, M., et al., Regulation of the CRL4(Cdt2) ubiquitin ligase and cell-cycle exit by the SCF(Fbxo11) ubiquitin ligase[J]. Mol Cell, 2013. 49(6): p. 1159-66.
[129] KUCHAY, S., et al., FBXL2- and PTPL1-mediated degradation of p110-free p85beta regulatory subunit controls the PI (3)K signalling cascade[J]. Nat Cell Biol, 2013. 15(5): p. 472-80.
[130] D'ANGIOLELLA, V., et al., Cyclin F-mediated degradation of ribonucleotide reductase M2 controls genome integrity and DNA repair[J]. Cell, 2012. 149(5): p. 1023-34.
[131] DUAN, S., et al., mTOR generates an auto-amplification loop by triggering the betaTrCP- and CK1alpha-dependent degradation of DEPTOR[J]. Mol Cell, 2011. 44(2): p. 317-24.
[132] ZHAO, Y., X. XIONG, AND Y. SUN, DEPTOR, an mTOR inhibitor, is a physiological substrate of SCF (betaTrCP) E3 ubiquitin ligase and regulates survival and autophagy[J]. Mol Cell, 2011. 44(2): p. 304-16.
[133] GAO, D., et al., mTOR drives its own activation via SCF (betaTrCP)-dependent degradation of the mTOR inhibitor DEPTOR[J]. Mol Cell, 2011. 44(2): p. 290-303.
[134] PUERTOLLANO, R., et al., The complex relationship between TFEB transcription factor phosphorylation and subcellular localization[J]. EMBO J, 2018. 37(11).
[135] BESSON, A., et al., A pathway in quiescent cells that controls p27Kip1 stability, subcellular localization, and tumor suppression[J]. Genes Dev, 2006. 20(1): p. 47-64.
[136] NAKAYAMA, K.I. AND K. NAKAYAMA, Ubiquitin ligases: cell-cycle control and cancer[J]. Nat Rev Cancer, 2006. 6(5): p. 369-81.
[137] NAKAYAMA, K.I. AND K. NAKAYAMA, Regulation of the cell cycle by SCF-type ubiquitin ligases[J]. Semin Cell Dev Biol, 2005. 16(3): p. 323-33.
[138] WU, Z.H., et al., Molecular linkage between the kinase ATM and NF-kappaB signaling in response to genotoxic stimuli[J]. Science, 2006. 311(5764): p. 1141-6.
[139] CHAPLAN, S.R., et al., Quantitative assessment of tactile allodynia in the rat paw[J]. J Neurosci Methods, 1994. 53(1): p. 55-63.
[140] HUANG, T., et al., Identifying the pathways required for coping behaviours associated with sustained pain[J]. Nature, 2019. 565(7737): p. 86-90.
[141] ROBERT M. GLEESON, M.G.D., NEIL F. KIRTON, Intracranial cannulation of small animals[J]. Behavior Research Methods & Instrumentation, 1980. 12(3): p. 346-348.
[142] BILLUPS, D. AND D. ATTWELL, Control of intracellular chloride concentration and GABA response polarity in rat retinal ON bipolar cells[J]. J Physiol, 2002. 545(1): p. 183-98.
[143] LANGSJO, J.W., et al., S-ketamine anesthesia increases cerebral blood flow in excess of the metabolic needs in humans[J]. Anesthesiology, 2005. 103(2): p. 258-68.
[144] GUO, Z.V., et al., Maintenance of persistent activity in a frontal thalamocortical loop[J]. Nature, 2017. 545(7653): p. 181-186.
[145] IACCARINO, H.F., et al., Gamma frequency entrainment attenuates amyloid load and modifies microglia[J]. Nature, 2016. 540(7632): p. 230-235.
[146] HEUBL, M., et al., GABAA receptor dependent synaptic inhibition rapidly tunes KCC2 activity via the Cl (-)-sensitive WNK1 kinase[J]. Nat Commun, 2017. 8(1): p. 1776.
[147] REITZ, S.L., et al., Activation of preoptic tachykinin 1 neurons promotes wakefulness oversleep and volatile anesthetic-induced unconsciousness[J]. Curr Biol, 2021. 31(2): p. 394-405 e4.
[148] ZHANG, Z., et al., Neuronal ensembles sufficient for recovery sleep and the sedative actions of alpha2 adrenergic agonists[J]. Nat Neurosci, 2015. 18(4): p. 553-561.
[149] BLAESSE, P., et al., Cation-chloride cotransporters and neuronal function[J]. Neuron, 2009. 61(6): p. 820-38.
[150] WILLIAMS, J.R. AND J.A. PAYNE, Cation transport by the neuronal K(+)-Cl(-) cotransporter KCC2: thermodynamics and kinetics of alternate transport modes[J]. Am J Physiol Cell Physiol, 2004. 287(4): p. C919-31.
[151] BLAESSE, P., et al., Oligomerization of KCC2 correlates with development of inhibitory neurotransmission[J]. J Neurosci, 2006. 26(41): p. 10407-19.
[152] CANCEDDA, L., et al., Excitatory GABA action is essential for morphological maturation of cortical neurons in vivo[J]. J Neurosci, 2007. 27(19): p. 5224-35.
[153] PUSKARJOV, M., et al., Activity-dependent cleavage of the K-Cl cotransporter KCC2 mediated by calcium-activated protease calpain[J]. J Neurosci, 2012. 32(33): p. 11356-64.
[154] ZHOU, H.Y., et al., N-methyl-D-aspartate receptor- and calpain-mediated proteolytic cleavage of K+-Cl- cotransporter-2 impairs spinal chloride homeostasis in neuropathic pain[J]. J Biol Chem, 2012. 287(40): p. 33853-64.
[155] DURING, M.J. AND D.D. SPENCER, Extracellular hippocampal glutamate and spontaneous seizure in the conscious human brain[J]. Lancet, 1993. 341(8861): p. 1607-10.
[156] BORETIUS, S., et al., Halogenated volatile anesthetics alter brain metabolism as revealed by proton magnetic resonance spectroscopy of mice in vivo[J]. Neuroimage, 2013. 69: p. 244-55.
[157] CHEN, X., et al., Glutamate diffusion in the rat brain in vivo under light and deep anesthesia conditions[J]. Magn Reson Med, 2019. 82(1): p. 84-94.
[158] MAHADEVAN, V., et al., Native KCC2 interactome reveals PACSIN1 as a critical regulator of synaptic inhibition[J]. Elife, 2017. 6.
[159] WOJCIK, C., VCP - the missing link in protein degradation? [J]. Trends Cell Biol, 2002. 12(5): p. 212.
[160] DOYON, N., et al., Efficacy of synaptic inhibition depends on multiple, dynamically interacting mechanisms implicated in chloride homeostasis[J]. PLoS Comput Biol, 2011. 7(9): p. e1002149.
[161] CONWAY, L.C., et al., N-Ethylmaleimide increases KCC2 cotransporter activity by modulating transporter phosphorylation[J]. J Biol Chem, 2017. 292(52): p. 21253-21263.
[162] ALAM, M.S., Proximity ligation assay (PLA)[J]. Curr Protoc Immunol, 2018. 123(1): p. e58.
[163] PADBERG, J. AND L. KRUBITZER, Thalamocortical connections of anterior and posterior parietal cortical areas in New World titi monkeys[J]. J Comp Neurol, 2006. 497(3): p. 416-35.
[164] HEMMINGS, H.C., JR., et al., Towards a comprehensive understanding of anesthetic mechanisms of action: a decade of discovery[J]. Trends Pharmacol Sci, 2019. 40(7): p. 464-481.
[165] RAVID, T. AND M. HOCHSTRASSER, Diversity of degradation signals in the ubiquitin-proteasome system[J]. Nat Rev Mol Cell Biol, 2008. 9(9): p. 679-90.
[166] RECHSTEINER, M. AND S.W. ROGERS, PEST sequences and regulation by proteolysis[J]. Trends Biochem Sci, 1996. 21(7): p. 267-71.
[167] ROGERS, S., R. WELLS, AND M. RECHSTEINER, Amino acid sequences common to rapidly degraded proteins: the PEST hypothesis[J]. Science, 1986. 234(4774): p. 364-8.
[168] RINEHART, J., et al., Sites of regulated phosphorylation that control K-Cl cotransporter activity[J]. Cell, 2009. 138(3): p. 525-36.
[169] HEMMINGS, H.C., JR., et al., Emerging molecular mechanisms of general anesthetic action[J]. Trends Pharmacol Sci, 2005. 26(10): p. 503-10.
[170] FRIEDEL, P., et al., WNK1-regulated inhibitory phosphorylation of the KCC2 cotransporter maintains the depolarizing action of GABA in immature neurons[J]. Sci Signal, 2015. 8(383): p. ra65.
[171] KAHLE, K.T., et al., WNK protein kinases modulate cellular Cl- flux by altering the phosphorylation state of the Na-K-Cl and K-Cl cotransporters[J]. Physiology (Bethesda), 2006. 21: p. 326-35.
[172] DE LOS HEROS, P., et al., The WNK-regulated SPAK/OSR1 kinases directly phosphorylate and inhibit the K+-Cl- co-transporters[J]. Biochem J, 2014. 458(3): p. 559-73.
[173] PIALA, A.T., et al., Chloride sensing by WNK1 involves inhibition of autophosphorylation[J]. Sci Signal, 2014. 7(324): p. ra41.
[174] CHEN, X., S. SHU, AND D.A. BAYLISS, HCN1 channel subunits are a molecular substrate for hypnotic actions of ketamine[J]. J Neurosci, 2009. 29(3): p. 600-9.
[175] PATEL, A.J., et al., Inhalational anesthetics activate two-pore-domain background K+ channels[J]. Nat Neurosci, 1999. 2(5): p. 422-6.
[176] PAVEL, M.A., et al., Studies on the mechanism of general anesthesia[J]. Proc Natl Acad Sci U S A, 2020. 117(24): p. 13757-13766.
[177] KELZ, M.B., et al., Escape from oblivion: neural mechanisms of emergence from general anesthesia[J]. Anesth Analg, 2019. 128(4): p. 726-736.
[178] YANARU, T., et al., Propofol-induced generalized tonic-clonic seizure: a case report[J]. Masui, 2010. 59(8): p. 1036-8.
[179] KIM, M.J., D.G. LIM, AND J.S. YEO, Refractory status epilepticus occurred at the end of sevoflurane anesthesia in patient with epilepsy[J]. Korean J Anesthesiol, 2013. 65(1): p. 93-4.
[180] TUNA, Y., A. TAS, AND S. KOKLU, Propofol-induced myoclonic seizures after endoscopic procedure in an elderly woman[J]. Chin Med J (Engl), 2012. 125(5): p. 785.
[181] KOFKE, W.A., Anesthetic management of the patient with epilepsy or prior seizures[J]. Curr Opin Anaesthesiol, 2010. 23(3): p. 391-9.
[182] VOSS, H.U., et al., Possible axonal regrowth in late recovery from the minimally conscious state[J]. J Clin Invest, 2006. 116(7): p. 2005-11.
[183] TATETSU, M., et al., GABA/glycine signaling during degeneration and regeneration of mouse hypoglossal nerves[J]. Brain Res, 2012. 1446: p. 22-33.
[184] KIM, J., et al., Changes in the expression and localization of signaling molecules in mouse facial motor neurons during regeneration of facial nerves[J]. J Chem Neuroanat, 2018. 88: p. 13-21.
[185] EWENS, C.A., et al., Structural and functional implications of phosphorylation and acetylation in the regulation of the AAA+ protein p97[J]. Biochem Cell Biol, 2010. 88(1): p. 41-8.
[186] HEIDELBERGER, J.B., et al., Proteomic profiling of VCP substrates links VCP to K6-linked ubiquitylation and c-Myc function[J]. EMBO Rep, 2018. 19(4).
[187] AHLSTEDT, B.A., R. GANJI, AND M. RAMAN, The functional importance of VCP to maintaining cellular protein homeostasis[J]. Biochem Soc Trans, 2022. 50(5): p. 1457-1469.
[188] BRAUN, R.J. AND H. ZISCHKA, Mechanisms of Cdc48/VCP-mediated cell death: from yeast apoptosis to human disease[J]. Biochim Biophys Acta, 2008. 1783(7): p. 1418-35.
[189] SCHUBERTH, C. AND A. BUCHBERGER, UBX domain proteins: major regulators of the AAA ATPase Cdc48/p97[J]. Cell Mol Life Sci, 2008. 65(15): p. 2360-71.
[190] MADSEN, L., et al., New ATPase regulators--p97 goes to the PUB[J]. Int J Biochem Cell Biol, 2009. 41(12): p. 2380-8.
[191] KAHLE, K.T., et al., Modulation of neuronal activity by phosphorylation of the K-Cl cotransporter KCC2[J]. Trends Neurosci, 2013. 36(12): p. 726-737.
[192] DE KONINCK, Y., Altered chloride homeostasis in neurological disorders: a new target[J]. Curr Opin Pharmacol, 2007. 7(1): p. 93-9.
[193] KAHLE, K.T., et al., Roles of the cation-chloride cotransporters in neurological disease[J]. Nat Clin Pract Neurol, 2008. 4(9): p. 490-503.
[194] LAUF, P.K. AND B.E. THEG, A chloride dependent K+ flux induced by N-ethylmaleimide in genetically low K+ sheep and goat erythrocytes[J]. Biochem Biophys Res Commun, 1980. 92(4): p. 1422-8.
[195] FUJISE, H. AND P.K. LAUF, Swelling, NEM, and A23187 activate Cl(-)-dependent K+ transport in high-K+ sheep red cells[J]. Am J Physiol, 1987. 252(2 Pt 1): p. C197-204.
[196] JENNINGS, M.L. AND R.K. SCHULZ, Okadaic acid inhibition of KCl cotransport. Evidence that protein dephosphorylation is necessary for activation of transport by either cell swelling or N-ethylmaleimide[J]. J Gen Physiol, 1991. 97(4): p. 799-817.