Binding of benzocaine in batrachotoxin-modified Na+ channels. State-dependent interactions.

Abstract

Hille (1977. Journal of General Physiology. 69:497-515) first proposed a modulated receptor hypothesis (MRH) to explain the action of benzocaine in voltage-gated Na+ channels. Using the MRH as a framework, we examined benzocaine binding in batrachotoxin (BTX)-modified Na+ channels under voltage-clamp conditions using either step or ramp command signals. We found that benzocaine binding is strongly voltage dependent. At -70 mV, the concentration of benzocaine that inhibits 50% of BTX-modified Na+ currents in GH3 cells (IC50) is 0.2 mM, whereas at +50 mV, the IC50 is 1.3 mM. Dose-response curves indicate that only one molecule of benzocaine is required to bind with one BTX-modified Na+ channel at -70 mV, whereas approximately two molecules are needed at +50 mV. Upon treatment with the inactivation modifier chloramine-T, the binding affinity of benzocaine is reduced significantly at -70 mV, probably as a result of the removal of the inactivated state of BTX-modified Na+ channels. The same treatment, however, enhances the binding affinity of cocaine near this voltage. External Na+ ions appear to have little effect on benzocaine binding, although they do affect cocaine binding. We conclude that two mechanisms underlie the action of local anesthetics in BTX-modified Na+ channels. Unlike open-channel blockers such as cocaine and bupivacaine, neutral benzocaine binds preferentially with BTX-modified Na+ channels in a closed state. Furthermore, benzocaine can be modified chemically so that it behaves like an open-channel blocker. This compound also elicits a use-dependent block in unmodified Na+ channels after repetitive depolarizations, whereas benzocaine does not. The implications of these findings for the MRH theory will be discussed.

Dopamine‐dependent and dopamine‐independent actions of cocaine as revealed by brain thermorecording in freely moving rats

“…Additionally, because a single cocaine injection induces a ∼26 µ m peak in brain concentration with a half‐time of decrease of about 10 min (Fowler et al ., 1998), brain levels of cocaine during self‐administration should be also much higher. At these higher levels, cocaine is highly effective as a Na + channel blocker (Gifford & Johnson, 1992), and its inhibiting effects are strongly amplified during depolarization and neuronal activation (Wang & Wang, 1994; Wright et al ., 1999; Kiyatkin & Rebec, 2000). Therefore, the relative efficiency of cocaine's action on DA uptake and Na + channels may show opposite changes with an increase in local cocaine levels, being gradually diminished and gradually enhanced, respectively.…”

Mechanism of sodium channel block by local anesthetics, antiarrhythmics, and anticonvulsants

“…Because this mechanism is obviously not applicable to neutral drugs, a model was proposed in which phenytoin, CMZ, and lamotrigine sterically occlude the pore (Lipkind and Fozzard, 2010). A small neutral benzocaine and its analogues, which do not fit any of these models, are proposed to block the channel by stabilizing the closed state (Wang and Wang, 1994;Wang et al, 1997). Such a diversity of mechanisms for the ligands that target the same region and display similar electrophysiological characteristics seems excessive.…”

Comment on Amézquita et al. (2017) “Conspicuousness, color resemblance, and toxicity in geographically diverging mimicry: The pan-Amazonian frogAllobates femoralis”

“…, ; Santos et al. ), and benzocaine interferes with Na + channels (Schneider and Dubois ; Wang and Wang ; Quan et al. ).…”

“…Benzocaine (10-20% solution) is a synthetic anesthetic commonly used for euthanizing frogs (McDiarmid 1994), typically by direct application to the skin where it is quickly absorbed; however, this poses a significant problem when studying the physiological and/or behavioral effects of skin secretions on other organisms. At the molecular level, frog alkaloids and benzocaine share similar modes of action: frog alkaloids interfere with various subtypes of nicotinic receptors and a diversity of ion channels (e.g., Na + , K + , Ca 2+ channels; Daly et al 1999Daly et al , 2005Santos et al 2016), and benzocaine interferes with Na + channels (Schneider and Dubois 1986;Wang and Wang 1994;Quan et al 1996). Also, benzocaine is known to inhibit locomotor activity in mice (Reith et al 1985) and cause methemoglobinemia (Davis et al 1993), the symptoms of which vary in humans from cyanosis, dizziness, lethargy, and syncope to seizures, arrhythmias, shock, and even death, depending on methemoglobin concentration (Rodriguez et al 1994).…”