Dr Mark Moore MD
Tallahassee's Anesthesiologist


The Molecular Biology of General Anesthesia

Reviews: What is consciousness? Is it something other than biological or physical? Is it purely physiological? Regardless of ones philosophical disposition on the “mind/body” issue, the short answer is that it must be in part physiological considering that it can be manipulated by purely physical means. Specifically, consciousness is manipulated thousands of times daily and nightly around the world in the use of general anesthesia. Interestingly, however, the molecular basis for these effects are not as well known as one might suspect considering the widespread use and relative safety of anesthetics.
In the early days of anesthetics, for a period of about 10 years from 1846 to 1946 three structurally diverse inhalation anesthetics were discovered and put to widespread use. These were chloroform, ether and nitrous oxide. The fascinating thing about these drugs is that they are structurally different from each other. Thus, the fundamental question in anesthesiology became, “how do volatile gases with divers structures give rise to the state of anesthesia?” (Mashour, 350)
To answer this fundamental question several hypothesis have been formulated. The first was the “unitary hypothesis” posited by Claude Bernard. He based this hypothesis on the idea that the anesthetics must have some common final mechanism since they all produced the same final effect: the state of anesthesia. Next came the “lipid hypothesis” with the independent discovery by Meyer and Overton that anesthetic potency was correlated with its solubility in oil. Thus, the lipid hypothesis proposed that anesthetics likely acted by perturbing lipid cell membranes of neural tissue. Then Franks and Lieb showed that a wide variety of anesthetics could inhibit the activity of a prepared lipid-free enzyme. This gave way to the “protein hypothesis” which posits that proteins could be the specific and direct targets of inhaled anesthetic gases. It has been shown since that volatile anesthetic gases act on many proteins such as serum proteins, ion channels, and even intracellular signaling molecules like protein kinase (Mashour, 350).
The general consensus is that ion channels provide the most likely target of anesthetic action. This is because ion channels have diverse roles in neural activity. They establish neuronal membrane potentials, propagate action potentials, moderate pre-synaptic release, and activate in response to post-synaptic binding. A few general types of ion channels are affected; ligand-gated ion channels, calcium channels, and potassium channels, both voltage gated and non-voltage gated (Mashour, 351).
Research is currently heavily directed towards gamma-aminobutyric acid type A receptors (GABAa), neuronal nicotinic acetylcholine receptors (nnAch), and Glutamate N-methyl-D-aspartate receptors, and glycine receptors in ligand-gated ion channels. One study examined chimeric glycine/p subunits and identified regions of the protein that contributed to the potentiating effects of ethanol and enflurane. Particularly Ser267 and Ala288. Other research has concluded that hydrophobic cavities with the presence of methionine and aromatic residues are in principal the ideal binding sites for anesthetics (Thompson, 79,80).
In addition, advances in neuroscience and technology give us greater detail and vision into the workings of active conscious and unconscious brains. Electro-Encephalograms and functional magnetic resonance imaging may lead to breakthroughs in the physiological and molecular foundations of consciousness, as well as genetic manipulation such as knock-out and “knock-in” mice that enable assessment of the roles of small regions within proteins (Mashour, 355).

Research: Inhalation general anesthetics have recently been shown to inhibit neuronal nicotinic acetylcholine (ACh) receptors (nnAChRs) expressed in Xenopus laevis oocytes and in molluscan neurons. (Mori, 732) However, these systems are not model systems for mammalian neurons. Thus the purpose of the experiment in question was to understand how volatile inhalation gaseous anesthetics might modulate neuronal nicotinic acetylcholine receptors in rat cortical neurons. Halothane was used as the anesthetic and its action on the nnAChRs in rat neurons in primary long-term culture was examined in detail. (Mori, 732)

The cells were prepared from fetuses removed from a 17-day pregnant Sprague-Dawley rat under methoxyflurane anesthesia. Small wedges of frontal cortex were excised and subsequently incubated in phosphate-buffered saline solution. The purpose was to form a cortical neuron/glia coculture of which cells that had cultured for 2 to 9 weeks were then used in the electrophysiological experiments. The concentrations of halothane used in the experiments were 7.5 to 2500 mM. The whole-cell, patch-clamp technique was used to record ionic currents induced by ACh application through a U-tube system. ACh and halothane were coapplied through a U-tube, and halothane was perfused through the bath starting 2 min before the coapplication. Two-minute preperfusion was long enough to exchange the whole bath solution and to allow halothane to exhibit its effect (Mori, 733). The results showed that halothane inhibited the currents induced by 300 mM ACh in a concentration-dependent manner. The inhibition was reversible after washout of halothane. (Mori, 735).

The study showed that halothane could reversibly inhibit a7-type and a4b2-type currents of nnAChRs in a concentration-dependent manner at clinically relevant concentrations. Halothane block of a7-type currents was independent of ACh concentration, whereas the block of a4b2-type currents became less with decreasing ACh
concentration. (Mori, 735)

Ultimately, the contribution this research makes is important since it establishes that nnAChRs can be inhibited in mammalian neuronal models with volatile gaseous anesthetics in a manner consisted with clinical usage. This is relevant and important since the primary issue in finding molecular mechanisms in the state of anesthesia may ultimately rest on protein interactions of this type with ligand specific inhibition and washout controlling the functional behaviors of consciousness and amnesia in humans.

Many things are still unknown about this process. For example, it is not known how consciousness is propagated within the central nervous system. This type of research, however suggests that physiological neural networks and their detailed molecular biology may lead to the elucidation of a mechanism, which is verifiable and testable. This is of primary importance in establishing that molecular biology and its physical entailments are ultimately the “stuff” of consciousness and rationality. The limitations however of this study is that it could only involve in vitro experimentation on the neural/glial cells and not in vivo.

1. G. Mashour, S. Forman, J. Campagna, Mechanisms of general anesthesia: from molecules to mind, 2005, Best Practice & Research Clinical Anaesthesiology, Vol. 19. No. 3, pp. 349-364
2. S. Thompson, K. Wafford, Mechanism of action of general anaesthetics- new information from molecular pharmacology, 2001, Current Opinion in Pharmacology, 1:78-83
3. Mori, Zhao, Zuo, Aistrup, Nishikawa, Marszalec,
Yeh, Narahashi, Modulation of Neuronal Nicotinic Acetylcholine Receptors by
Halothane in Rat Cortical Neurons, 2001, Molecular Pharmacology 59:732-743

Ray S. Magill - 11/16/05

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