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Department of Biology, Williams College, Williamstown, Massachusetts 01267
Ethyl-m-aminobenzoate, known as tricaine, metacaine, or MS-222, is a commonly used general anesthetic for fish and amphibia. This anesthetic depresses neuronal activity in both the peripheral (1) and central nervous systems (2) of fish, but its effects on the basic physiological properties of individual neurons are unknown. Clearly, the action of tricaine must be determined, if neurophysiological data obtained during its use are to be correctly interpreted. We have therefore studied the effect of tricaine on the level of current necessary to elicit an action potential and spike height of supramedullary/dorsal neurons of the cunner Tautogolabrus adspersus (3,4).
Responses of supramedullary/dorsal cells to depolarizing current pulses were recorded from cunner, 8–13.9 cm in body length; the fish were either under general or local anesthesia. In the first condition (general anesthesia), eight fish were initially anesthetized in tricaine (300 mg/l, Sigma) in seawater with sodium bicarbonate (450 mg/l) added. When respiration ceased, the fish were transferred to an operating chamber where tricaine (100 mg/l) in chilled seawater with sodium bicarbonate (450 mg/l; pH = 7.2) was recirculated through the mouth and over the gills for the duration of the experiment. In the second condition (local anesthesia), 10 fish were injected with pancuronium bromide (0.1 mg/kg; Sigma) to block neuromuscular transmission and were then placed in an operating chamber, where a respiratory current of chilled seawater was applied through the mouth and over the gills. Local anesthetic (20% benzocaine gel, Ultradent) was applied to the skin overlying the rostral spinal cord. After 5 min, the skin was removed and local anesthetic was applied to the underlying muscle. After 5 min, the rostral spinal cord was exposed. Single microelectrode recordings (3 M KCl filled, 5–15 M
) were then made from supramedullary/dorsal cell somata.
Under general anesthesia, action potentials 77 ± 8 mV (mean ± SD, n = 12) in amplitude could be evoked by 7 ± 2 nA of current. Under local anesthesia, spike height (91 ± 6 mV, n = 12) was significantly higher (P < 0.001; unpaired t test) and the current needed to evoke the spike was significantly lower (3 ± 2 nA; P < 0.001; unpaired t test) than values obtained under general anesthesia. These changes occurred without significant shifts in resting membrane potential (general anesthesia = -68 ± 5 mV; local anesthesia = -66 ± 5 mV).
Recordings were made in the supramedullary/dorsal cell soma, a location that is believed to be distant from the spike initiating zone (5). If the position of the microelectrode varied in its proximity to the spike initiating zone between experiments, then one might expect anesthetic-independent variation in spike height and current needed to elicit the spike. We controlled for this possibility in three experiments by continuously monitoring supramedullary/dorsal cells without moving the microelectode while the levels of tricaine were changed. The spike height decreased and current needed to elicit the spike increased upon the addition of tricaine (100 mg/l) to the water (Fig. 1). When the tricaine was removed by running anesthetic-free seawater over the gills, the current needed returned close to pre-anesthetic levels, and the spike height began to recover.
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The physiological effects of tricaine on individual supramedullary/dorsal neurons support previous findings that this anesthetic is blocking sodium channels. Specifically, voltage clamp studies on the squid giant axon have shown that the internal application of tricaine caused a major reduction of the peak sodium currents and a smaller reduction of potassium currents; these effects were reversible (7).
This study indicates that tricaine significantly alters afferent input, spike height, and current needed to elicit an action potential of supramedullary/dorsal cells. Consequently, physiological measurements must be viewed with caution if tricaine is used as a general anesthetic. The physiological effects of tricaine on supramedullary/dorsal cells are partially reversible, but the possibility that this anesthetic has long-term residual effects remains to be determined.
This work was supported in part by Howard Hughes Medical Institute and Essel Foundation grants to Williams College.
Literature Cited
This article has been cited by other articles:
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  S. J. Zottoli, O. T. Burton, J. A. Chambers, R. Eseh, L. M. Gutierrez, and M. M. Kron Transient Use of Tricaine to Remove the Telencephalon Has No Residual Effects on Physiological Recordings of Supramedullary/Dorsal Neurons of the Cunner, Tautogolabrus adspersus Biol. Bull., October 1, 2003; 205(2): 211 - 212. [Full Text] [PDF]
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