Findings produced by a team of researchers from the Hemmings Lab and across Weill Cornell Medical College were published electronically in the Early Edition of the journal, Proceedings of the National Academy of Sciences (PNAS).
Joel P. Baumgarta,1, Zhen-Yu Zhoua,1, Masato Haraa,b, Daniel C. Cooka, Michael B. Hoppac,d, Timothy A. Ryana,c, and Hugh C. Hemmings Jr.a,e,2 Isoflurane inhibits synaptic vesicle exocytosis through reduced Ca2+ influx, not Ca2+-exocytosis coupling. PNAS. Early edition. 12 Aug 2015. www.pnas.org/cgi/doi/10.1073/pnas.1500525112
Department of Anesthesiology, Weill Cornell Medical College, New York, NY 10065; bDepartment of Anesthesiology, Kurume University School of Medicine, Kurume, Fukuoka 830-0011, Japan; cDepartment of Biochemistry, Weill Cornell Medical College, New York, NY 10065; dDepartment of Biology, Dartmouth College, Hanover, NH 03755; and eDepartment of Pharmacology, Weill Cornell Medical College, New York, NY 10065
“Overview: Identifying presynaptic mechanisms of general anesthetics is critical to understanding their effects on synaptic transmission. We show that the volatile anesthetic isoflurane inhibits synaptic vesicle (SV) exocytosis at nerve terminals in dissociated rat hippocampal neurons through inhibition of presynaptic Ca2+ influx without significantly altering the Ca2+ sensitivity of SV exocytosis. A clinically relevant concentration of isoflurane (0.7 mM) inhibited changes in [Ca2+]i driven by single action potentials (APs) by 25 ±3%, which in turn led to 62 ± 3% inhibition of single AP-triggered exocytosis at 4 mM extracellular Ca2+ ([Ca2+]e). Lowering external Ca2+ to match the isoflurane-induced reduction in Ca2+ entry led to an equivalent reduction in exocytosis. These data thus indicate that anesthetic inhibition of neurotransmitter release from small SVs occurs primarily through reduced axon terminal Ca2+ entry without significant direct effects on Ca2+-exocytosis coupling or on the SV fusion machinery. Isoflurane inhibition of exocytosis and Ca2+ influx was greater in glutamatergic compared with GABAergic nerve terminals, consistent with selective inhibition of excitatory synaptic transmission. Such alteration in the balance of excitatory to inhibitory transmission could mediate reduced neuronal interactions and network-selective effects observed in the anesthetized central nervous system.
Significance: Clarification of the presynaptic actions of general anesthetics is critical to understanding the molecular and cellular mechanisms of their prominent effects on synaptic transmission. We show that the ether anesthetic isoflurane inhibits synaptic vesicle exocytosis through inhibition of presynaptic Ca2+ influx in the absence of significant alteration of the Ca2+ sensitivity of exocytosis. The greater inhibition of glutamate release compared with GABA release is explained by the relative anesthetic resistance of Ca2+ influx in GABAergic boutons, consistent with overall reduction in excitatory synaptic tone."
Mattusch C1, Kratzer S, Buerge M, Kreuzer M, Engel T, Kopp C, Biel M, Hammelmann V, Ying SW, Goldstein PA, Kochs E, Haseneder R, Rammes G. Impact of Hyperpolarization-activated, Cyclic Nucleotide-gated Cation Channel Type 2 for the Xenon-mediated Anesthetic Effect: Evidence from In Vitro and In Vivo Experiments. Anesthesiology. 2015 May;122(5):1047-59.
1From the Department of Anesthesiology, Klinikum rechts der Isar, Technische Universität München, Munich, Germany (C.M., S.K., M. Buerge, M.K., T.E., C.K., E.K., R.H., G.R.); Department Pharmazie-Zentrum für Pharmaforschung, Ludwig-Maximilians-Universität München, Munich, Germany (M. Biel, V.H.); and Department of Anesthesiology, Weill Cornell Medical College, New York, New York (S.-W.Y., P.A.G.).
'Background: The thalamus is thought to be crucially involved in the anesthetic state. Here, we investigated the effect of the inhaled anesthetic xenon on stimulus-evoked thalamocortical network activity and on excitability of thalamocortical neurons. Because hyperpolarization-activated, cyclic nucleotide-gated cation (HCN) channels are key regulators of neuronal excitability in the thalamus, the effect of xenon on HCN channels was examined.
Methods: The effects of xenon on thalamocortical network activity were investigated in acutely prepared brain slices from adult wild-type and HCN2 knockout mice by means of voltage-sensitive dye imaging. The influence of xenon on single-cell excitability in brain slices was investigated using the whole-cell patch-clamp technique. Effects of xenon on HCN channels were verified in human embryonic kidney cells expressing HCN2 channels.
Results: Xenon concentration-dependently diminished thalamocortical signal propagation. In neurons, xenon reduced HCN channel-mediated Ih current amplitude by 33.4 ± 12.2% (at −133 mV; n = 7; P = 0.041) and caused a left-shift in the voltage of half-maximum activation (V1/2) from −98.8 ± 1.6 to −108.0 ± 4.2 mV (n = 8; P = 0.035). Similar effects were seen in human embryonic kidney cells. The impairment of HCN channel function was negligible when intracellular cyclic adenosine monophosphate level was increased. Using HCN2−/− mice, we could demonstrate that xenon did neither attenuate in vitro thalamocortical signal propagation nor did it show sedating effects in vivo.
Conclusions: Here, we clearly showed that xenon impairs HCN2 channel function, and this impairment is dependent on intracellular cyclic adenosine monophosphate levels. We provide evidence that this effect reduces thalamocortical signal propagation and probably contributes to the hypnotic properties of xenon.'
Paul M. Heerdt, MD, PhD, was part of a research team, which included researchers from Weill Cornell Medical College and the Cornell University College of Veterinary Medicine, whose findings were published in the American Journal of Veterinary Research in March. Dr. Heerdt's work was part of a growing collaboration between Weill Cornell and the College of Verterinary Medicine in Ithaca, which included the Residency Program's first rotation in Veterniary Medicine.
Manuel Martin-Flores, DVM; Jonathan Cheetham, VetMB, PhD; Luis Campoy, LV; Daniel M. Sakai, DVM; Paul M. Heerdt MD, PhD; Robin D. Gleed, BVSc. AJVR. Effect of gantacurium on evoked laryngospasm and duration of apnea in anesthetized healthy cats. 2015 March; 76 (3): 216-23.
"Objective: To evaluate whether the ultrashort-acting neuromuscular blocking agent gantacurium can be used to blunt evoked laryngospasm in anesthetized cats and to determine the duration of apnea without hemoglobin desaturation.
Animals: 8 healthy adult domestic shorthair cats.
Procedures: Each cat was anesthetized with dexmedetomidine and propofol, instrumented with a laryngeal mask, and allowed to breathe spontaneously (fraction of inspired oxygen, 1.0). The larynx was stimulated by spraying sterile water (0.3 mL) at the rima glottidis; a fiberscope placed in the laryngeal mask airway was used to detect evoked laryngospasm. Laryngeal stimulation was performed at baseline; after IV administration of gantacurium at doses of 0.1, 0.3, and 0.5 mg/kg; and after the effects of the last dose of gantacurium had terminated. Duration of apnea and hemoglobin oxygen saturation (measured by means of pulse oximetry) after each laryngeal stimulation were recorded. Neuromuscular block was monitored throughout the experiment by means of acceleromyography on a pelvic limb.
Conclusions and Clinical Relevance: Gantacurium may reduce tracheal intubation-associated morbidity in cats breathing oxygen."
Karl Herold, MD, PhD, Hugh C. Hemmings, Jr., MD, PhD, the Hemmings Lab, and others from Weill Cornell Medical College, had research findings published in the Journal of General Physiology in November.
Karl F. Herold,1 R. Lea Sanford,2 William Lee,1 Margaret F. Schultz,1 Helgi I. Ingólfsson,3 Olaf S. Andersen,2 and Hugh C. Hemmings Jr.1,3. Volatile anesthetics inhibit sodium channels without altering bulk lipid bilayer properties. J Gen Physiol. 2014 Dec;144(6):545-60.
1Department of Anesthesiology,2Department of Physiology and Biophysics, and 3Department of Pharmacology, Weill Cornell Medical College, New York, NY 10065
"Although general anesthetics are clinically important and widely used, their molecular mechanisms of action remain poorly understood. Volatile anesthetics such as isoflurane (ISO) are thought to alter neuronal function by depressing excitatory and facilitating inhibitory neurotransmission through direct interactions with specific protein targets, including voltage-gated sodium channels (Na(v)). Many anesthetics alter lipid bilayer properties, suggesting that ion channel function might also be altered indirectly through effects on the lipid bilayer. We compared the effects of ISO and of a series of fluorobenzene (FB) model volatile anesthetics on Na(v) function and lipid bilayer properties. We examined the effects of these agents on Na(v) in neuronal cells using whole-cell electrophysiology, and on lipid bilayer properties using a gramicidin-based fluorescence assay, which is a functional assay for detecting changes in lipid bilayer properties sensed by a bilayer-spanning ion channel. At clinically relevant concentrations (defined by the minimum alveolar concentration), both the FBs and ISO produced prepulse-dependent inhibition of Na(v) and shifted the voltage dependence of inactivation toward more hyperpolarized potentials without affecting lipid bilayer properties, as sensed by gramicidin channels. Only at supra-anesthetic (toxic) concentrations did ISO alter lipid bilayer properties. These results suggest that clinically relevant concentrations of volatile anesthetics alter Na(v) function through direct interactions with the channel protein with little, if any, contribution from changes in bulk lipid bilayer properties. Our findings further suggest that changes in lipid bilayer properties are not involved in clinical anesthesia."