Abstract
This study compares the actions of the intravenous anaesthetics propofol and ketamine on animal behaviour and neuronal activity in the snail Lymnaea stagnalis, particularly in relation to excitatory effects observed clinically. When injected into the whole animal, neither agent induced total anaesthesia. Rather, behavioural activity was enhanced by propofol (10−5 M) and ketamine (10−7 M), indicating excitatory effects. When superfused over the isolated central nervous system (CNS), differential effects were produced in two identified neurons, right pedal dorsal 1 (RPeD1) and visceral dorsal 4 (VD4). Resting membrane properties were largely unaffected. However, spike after hyperpolarisation was significantly reduced in RPeD1, but not VD4, with some evidence of increased excitability. In addition, an intrinsic bursting property (post-stimulus burst) in VD4 was altered by propofol (10−7 M). The results suggest significant excitatory components in the actions of some intravenous anaesthetics, as well as a potential role in modifying excitation and bursting mechanisms in the CNS.
Introduction
The intravenous anaesthetics, propofol and ketamine are used alone, or in conjunction with other agents, to induce or maintain anaesthesia. In addition to their anaesthetic effects, both propofol and ketamine have been shown clinically to induce excitatory (seizure-like) episodes (Kayama and Iwama, 1972, Winter, 1972, Sneyd, 1992). With propofol, these episodes are usually observed at low concentrations or during recovery where it could be assumed that the concentration of anaesthetic is low (Sutherland and Burt, 1994). The mechanism(s) underlying excitation is unknown, but it clearly has implications for the use of both agents in a clinical setting.
In vitro studies have shown different mechanisms of action for propofol and ketamine at the molecular level. Propofol’s main action appears to be as a GABA-A receptor modulator, enhancing GABA-mediated inhibition (Hara et al., 1993). Ketamine is primarily a non-competitive NMDA receptor antagonist, blocking the open receptor at a site within the pore (Orser et al., 1997). In addition, other cellular actions, including effects on nicotinic acetylcholine and 5-HT3 receptors (Violet et al., 1997) and ion channels and conductances (Yamakage et al., 1995, Wu et al., 1997) have been reported for both agents, but these do not explain the excitatory effects described above. A better knowledge of the cellular modes of action of these agents would further our understanding of their anaesthetic potential and that of related compounds, particularly in relation to minimising their excitatory actions. This study aimed to compare and contrast the effects of ketamine and propofol on specific, identified, neurons, using relevant concentrations in a model organism, the freshwater gastropod mollusc, Lymnaea stagnalis.
Molluscan neurons have been widely used for investigating and screening the effects of pharmacological agents (Girdlestone et al., 1989a, Girdlestone et al., 1989b, Mills et al., 1992). This is largely due to their highly accessible central nervous systems, with ‘giant’ neurons which can be individually identified from one preparation to the next (Kyriakides et al., 1989). This means that the same neuron, with known characteristics, as well as identified synaptic connections, can be used to study and compare responses to different agents within an intact nervous system in vitro. L. stagnalis has been exploited in studies of the actions of inhalation anaesthetics such as halothane, isoflurane and enflurane. All of these agents induce full and reversible anaesthesia in the whole animal, measured as a loss of the whole body withdrawal reflex, at clinically relevant doses, with an ED50 close to the anaesthetic requirement of mammals (Girdlestone et al., 1989a). The effects of halothane at the cellular level in this species have also been investigated. Halothane (1%) abolished excitatory, but not inhibitory, synaptic potentials between identified neurons; however, inhibitory inputs were blocked by 2% halothane (Spencer et al., 1996). Franks and Lieb (1988) described a novel K+ current (Kan) in neurons of L. stagnalis, which was activated by volatile anaesthetics, leading to hyperpolarisation. Studies such as these illustrate the potential of this organism for anaesthetic research.
In this study we examined the effects of propofol and ketamine on identified neurons, right pedal dorsal 1 (RPeD1) and visceral dorsal 4 (VD4) in the central ganglia of L. stagnalis. These neurones are known to form part of a central pattern-generating network that controls rhythmic respiratory movements for ventilation of the lung in this pulmonate mollusc (Syed et al., 1990, Syed and Winlow, 1991). In addition, a behavioural study assessed the anaesthetic potential of propofol and ketamine in the snail. The invertebrate anaesthetic menthol (Haydon et al., 1982) was also used for comparative purposes, allowing us to compare the behavioural and cellular effects of propofol and ketamine with those of an agent which is known to induce whole body anaesthesia in L. stagnalis (Girdlestone et al., 1989a).
Section snippets
Materials and methods
L. stagnalis (mean shell length 25 mm) were obtained from suppliers (Blades, Kent) and kept in aerated tanks of standard snail water (SSW, composition (mM); CaCl2, 2.0; MgSO4, 0.08; KHCO3, 0.04; KNO3, 0.05; NaHCO3, 0.07; pH 6.8–7.2) for at least 2 weeks prior to experimentation. For behavioural experiments, snails were starved for 48 h prior to the experiment, so as to minimise individual differences in motivational state (Tuersley and McCrohan, 1987).
Behaviour
Snails injected with propofol, Intralipid or snail saline showed an initially low BSS 2 min p
