Actions Of EtOH Research Articles

Double effect of ethanol on intracellular Ca 2+ levels in undifferentiated PC12 cells

In PC12, a cellular line derived from a rat pheochromocytoma, ethanol (EtOH) induces a different effect depending on the concentration used. When resting cells are incubated with an alcohol concentration less than or equal to 120 mM, the [Ca 2+] i increased with a double phase pattern. If the alcohol concentration was increased over 120–160 mM, EtOH reversed its effect and the [Ca 2+] i decreased. This decrease was strongly inhibited if KCl-depolarized cells were used and was completely abolished if the substrate constituted EtOH-chronically treated cells. The Ca 2+ increase is the consequence of an activation of L-type voltage-activated channels, while the other voltage-dependent channels (N-type), the receptor-operated channels and the Ca 2+ extrusion pump present in these cells are not involved in EtOH action. These findings indicate that EtOH can induce (by different mechanisms) both potentiating and inhibiting effects on [Ca 2+] i in PC12 cells in relation to the alcohol dose effectively present in the suspension medium.

Effects of prenatal ethanol exposure on later sensitivity to the low-dose stimulant actions of ethanol in mouse offspring: possible role of catecholamines.

The purpose of this study was to examine whether prenatal ethanol (EtOH) exposure alters later sensitivity to the low-dose stimulant effects of EtOH. Because the locomotor stimulant effects of EtOH are thought to be mediated, at least in part, by activation of brain monoamine systems, and because prenatal EtOH exposure has been shown to alter brain monoamine activity, it was hypothesized that prenatal EtOH exposure may alter sensitivity to the stimulant actions of EtOH. To test this hypothesis, sensitivity to the locomotor stimulant effects of various challenge doses of EtOH was examined in male and female offspring from prenatal alcohol (A), pair-fed (PF), and lab chow (LC) groups at different ages. In addition, to address the hypothesis further, sensitivity to the catecholamine synthesis inhibitor alpha-methyl-p-tyrosine (AMPT) was examined in these offspring, as well. Results indicated that male offspring prenatally exposed to EtOH exhibited reduced baseline activity and a blunted stimulant response to all challenge doses of EtOH (0.75-1.5 g/kg) in comparison with control offspring at 30 days of age, but these effects appeared to "normalize" at 70 days of age. Female EtOH-exposed offspring also exhibited a reduced baseline level of activity relative to control offspring, as well as a blunted stimulant response to the lowest challenge dose of EtOH (0.75 g/kg) at 30 days of age, and these effects persisted into adulthood. The stimulant response to higher doses of EtOH did not significantly differ among prenatal treatment groups in young or adult female offspring. However, because baseline activity was significantly lower in female EtOH-exposed offspring than control offspring, the stimulant response to these doses of EtOH (1.125 and 1.5 g/kg) was relatively greater than that for PF and LC offspring. Importantly, none of the differences in performance among the prenatal treatment groups could be attributed to an alteration in EtOH pharmacokinetics, because blood EtOH levels measured immediately following the 10-min test session were similar for all prenatal treatment groups across all of the EtOH test doses. Further, a similar response profile as that observed following EtOH challenge at 70 days of age was obtained following phenobarbital challenge (10-40 mg/kg). Finally, whereas AMPT (50-400 mg/kg) dose-dependently antagonized the stimulant effects of EtOH in all prenatal treatment groups, this effect of AMPT was significantly greater in mice prenatally exposed to EtOH in comparison with control offspring.(ABSTRACT TRUNCATED AT 400 WORDS)

MONITORING OF ETHANOL LEVELS IN THE RAT NUCLEUS ACCUMBENS BY BRAIN MICRODIALYSIS

Brain microdialysis was used to measure nucleus accumbens (NAC) ethanol (EtOH) over several hours after intraperitoneal (i.p.) injection of EtOH. Levels of EtOH in NAC perfusates were assayed by gas chromatography. Peak EtOH levels were observed 40 min after the injection and the three doses of EtOH, 0.5, 1.0, and 2.0 g/kg, resulted in peak perfusate levels of 5 mM, 18 mM, and 38 mM, respectively. Measurement of regional brain EtOH levels may prove useful for studying the relationship between patterns of alcohol consumption and the action of EtOH in the central nervous system.

Ca(2+)-independent and Ca(2+)-dependent stimulation of quantal neurosecretion in avian ciliary ganglion neurons.

1. Although it is generally agreed that Ca2+ couples depolarization to the release of neurotransmitters, hypertonic saline and ethanol (ETOH) evoke neurosecretion independent of extracellular Ca2+. One possible explanation is that these agents release Ca2+ from an intracellular store that then stimulates Ca(2+)-dependent neurosecretion. An alternative explanation is that these agents act independently of Ca2+. 2. This work extends previous observations on the action of ETOH and hypertonic solutions (HOSM) on neurons to include effects on [Ca2+]i. We have looked for Ca(2+)-independent or -dependent neurosecretion evoked by these agents in parasympathetic postganglionic neurons dissociated from chick ciliary ganglia and maintained in tissue culture. The change in concentration of free Ca2+ in the micromolar range inside neurons ([Ca2+]i) was measured with indo-1 with the use of a Meridian ACAS 470 laser scanning microspectrophotometer. 3. Elevated concentration of extracellular KCl increased [Ca2+]i and the frequency of quantal events. Also, a twofold increase in osmotic pressure (HOSM) produced a similar increase in quantal release and a significant rise in [Ca2+]i; however, the Ca2+ appeared to come from intracellular stores. 4. In contrast, ETOH stimulated quantal neurosecretion without a measurable change in [Ca2+]i. It appears the alcohol exerts its influence on some stage in the process of exocytosis that is distal to or independent of the site of Ca2+ action. 5. The effects of high [KCl]o and osmotic pressure were occlusive. This is explained in part by the observation that hypertonicity reduced Ca2+ current, but an action on Ca2+ stores is also likely.(ABSTRACT TRUNCATED AT 250 WORDS)

The involvement of NMDA receptors in acute and chronic effects of ethanol.

Recent evidence indicates involvement of excitatory amino acid receptors sensitive to N-methyl-d-aspartate (NMDA) in the action of ethanol (EtOH). Pronounced inhibition of NMDA receptor function is seen in vitro with concentrations of EtOH corresponding to those present during alcohol intoxication in humans. The present study was devoted to investigate the role of NMDA receptors in the action of EtOH in rats. Acute experiments showed antagonism by EtOH of convulsions induced by intracerebroventricular injection of NMDA. A similar effect was seen with a high dose of diazepam. Convulsions induced by an agonist of another excitatory amino acid receptor subtype, kainate, were also inhibited by EtOH. An uncompetitive antagonist of NMDA receptors, 5-methyl-10,11-dihydro-5H-dibenzocyclohepten-5,10-imine maleate (MK-801), potentiated EtOH-induced loss of righting, but attenuated the hypothermic action of EtOH. Moreover, MK-801 inhibited audiogenic convulsions in EtOH withdrawn rats. At the same time the effect of a proconvulsive dose of NMDA was not enhanced. Tolerance to the myorelaxant action of both EtOH and MK-801 upon repetitive administration was seen. Also some degree of cross-tolerance was observed. Moreover, MK-801 failed to modify EtOH preference in rats. The present results support involvement of NMDA receptors in expression of some acute and subchronic actions of EtOH and in expression of EtOH withdrawal.

Failure of Ro15-4513 to alter an ethanol-induced taste aversion

The ability of Ro15-4513, an imidazobenzodiazepine inverse benzodiazepine agonist, to attenuate/block the acquisition of an ethanol (ETOH)-induced conditioned taste aversion (CTA) was investigated in two experiments. Experiment 1 examined the effects of Ro15-4513 (3 mg/kg) on rats' consumption of a novel saccharin solution under a traditional CTA paradigm. Experiment 2 examined the effects of Ro15-4513 (3 mg/kg) on rats' consumption of a novel saccharin solution under a preexposure CTA paradigm. Under the preexposure paradigm, rats were given Ro15-4513 immediately before each of five daily consecutive preexposure treatments prior to the initial conditioning day. To obtain maximal preexposure and unconditioned stimulus effects, a 2-g/kg dose of ETOH (20% v/v) was used in the present study. As previously reported, animals given ETOH following 20-min access to a novel saccharin solution established moderate to strong aversions, with the degree of aversion being directly related to the number of conditioning days. Experiment 1 showed that Ro15-4513 failed to alter the CTA induced by ETOH. Experiment 2 further showed that Ro15-4513 failed to block the preexposure effect exerted on the ETOH-mediated CTA. The results confirm previous reports regarding the failure of Ro15-4513 to disrupt an ETOH-induced CTA. These data are in agreement with a number of behavioral studies demonstrating the failure of Ro15-4513 to antagonize certain actions of ETOH. Moreover, the present study along with a previous report suggests that ETOH-induced CTA's do not appear to be mediated via actions at the GABA-BDZ receptor complex.

Ethanol reduces vasopressin release by inhibiting calcium currents in nerve terminals

Ingestion of ethanol (EtOH) is known to result in a reduction of plasma arginine-vasopressin (AVP) levels in mammals. We examined the basis for this effect using a combination of biochemical and electrophysiological techniques. Release of AVP from nerve terminals isolated from the rat neurohypophysis was very sensitive to EtOH, with significant reductions in AVP release evident in 10 mM EtOH. However, EtOH did not affect the release of AVP from terminals which had been permeabilized with digitonin, suggesting that voltage-gated calcium channels might be the target of EtOH's actions. Patch clamping of these terminals indicated that both inactivating and long-lasting calcium currents were reduced in EtOH, but the long-lasting currents were more sensitive (significant reductions in 10 mM EtOH). EtOH-induced decreases in plasma AVP levels can be explained by EtOH's inhibition of calcium currents in the nerve terminals.

Ethanol increases the firing rate of dopamine neurons of the rat ventral tegmental area in vitro

The ventral tegmental area (VTA) is a brain region rich in dopamine-containing neurons. Since most agents which act as substrates for self-administration increase dopaminergic outflow in the mesolimbic or mesocortical areas, the VTA slice preparation may be useful for identifying drugs with potential for abuse. While ethanol (EtOH) is a drug of abuse which has been widely studied, the properties of ethanol which contribute to its abuse potential are not known. We have developed a brain slice preparation of the VTA in order to study the action of EtOH on putative dopamine neurons. Concentrations of EtOH from 20 to 320 mM produce a dose-dependent excitation of the dopamine-type neurons of the VTA. About 89% of neurons which have electrophysiological characteristics established for presumed dopamine-containing neurons were excited by ethanol in the pharmacologically relevant concentration range. This excitation persists in low-calcium, high-magnesium medium, which suggests a direct excitatory action of EtOH on dopamine-type cells in the VTA slice.

Ethanol and diazepam interactions on conflict behavior in rats

Twenty-four adult male rats trained on the Geller-Seifter conflict procedure received: 1.0, 2.0, or 4.0 mg/kg diazepam at 15 min, 0.25, 0.5, 1.0, 2.0, or 4.0 g/kg of ethanol (EtOH) at 30 min prior to the test session, or combinations of the same diazepam doses and 0.5 g/kg of EtOH. Ethanol administration produced a significant ( p<0.05) dose-related decrease in lever pressing during the periods prior to the introduction of the tone stimulus, compared to lever pressing during the corresponding control periods on the day prior to drug treatment. During the posttone periods 0.25 g/kg increased lever pressing ( p<0.05), while doses of EtOH from 0.5 to 4.0 g/kg resulted in significantly lower levels of lever pressing in periods following the discontinuation of the tone. In the presence of the conflict evoking tone stimulus, rats treated with 2.0 g/kg of EtOH increased their rate of lever pressing, while animals receiving the other EtOH doses exhibited lever pressing comparable to that observed during the corresponding control periods. Administration of EtOH had no effect on the number of reinforcements received during the entire test session, except for a significant ( p<0.05) reduction at the highest dose of 4.0 g/kg. Diazepam increased the number of reinforcements received during the entire test session as compared to those obtained during control sessions. Lever pressing was not affected by diazepam treatment during pretone or tone periods. During the posttone periods responses were significantly ( p<0.05) increased in rats receiving the lowest dose of 1.0 mg/kg of diazepam but the other doses were without effect. Combined exposure to 0.5 g/kg of EtOH and either 1.0, 2.0 or 4.0 mg/kg of diazepam resulted in significant ( p<0.05) reductions in lever pressing during the pretone periods compared to that observed with diazepam alone. The combination of EtOH with these doses of diazepam had no further effect on lever pressing during the posttone periods, or on the number of reinforcements received during the entire test session, compared to effects observed with diazepam alone. However, a significant ( p<0.01) interactive effect of EtOH and diazepam was observed for lever pressing in the presence of the conflict evoking tone stimulus. Exposure to 4.0 mg/kg of diazepam and 0.5 g/kg EtOH resulted in a marked increase in lever pressing in the presence of the tone stimulus, whereas either of these drug doses alone, as well as combinations of other diazepam doses and EtOH, had no effect on lever pressing during the tone periods. These results support an anxiolytic action of EtOH, defined as an increase in punished responding; as well as a sedative action, defined as a decrease in unpunished responding. In addition, diazepam potentiates anxiolytic and sedative actions of EtOH.

Ethanol suppression of schedule-controlled responding: Interactions with Ro 15-4513, Ro 15-1788 and CGS 8216

Rats (N = 14) were trained to respond under a five seconds differential reinforcement of low rate (DRL 5″) schedule and under a fixed ratio 10 (FR10) schedule of reinforcement. Ro 15-1788 did not influence the number of responses in the DRL 5″ schedule, but increased responding in the FR10 schedule. Ethanol (ETOH, 1250 mg/kg) and CGS 8216 (5 mg/kg) suppressed responding in both schedules and these effects were not antagonized by Ro 15-1788. The response suppressing effects of ETOH in both schedules were not influenced by CGS 8216. These results indicate that the response suppressing effects of ETOH and CGS 8216 are not mediated by the BDZ receptor. Ro 15-4513 suppressed responding strongly in the FR10 schedule. The response suppressing effects of Ro 15-4513 were additive with the response suppressing effects of ETOH. In rats (N=11) trained to respond under a variable interval 40 seconds-fixed ratio 10 (VI 40″-FR10) schedule Ro 15-4513 dose-dependently suppressed responding. These results indicate that Ro 15-4513 has sedative effects and is not able to antagonize all the behavioral actions of ETOH.

Alkanol effects on early potassium currents in Aplysia neurons depend on chain length.

The relationship between alkanol chain length and effects on the transient potassium current, IA, were examined in three identified Aplysia californica neurons with ethanol (EtOH), butanol (BuOH), and hexanol (HxOH). Qualitative differences were found when the actions of EtOH were compared with those of the longer-chain-length alcohols. Whereas EtOH primarily affected the decay time constant of IA, having minimal effects on amplitude, BuOH and HxOH exerted their major effect on the amplitude of IA, reducing it, while their effects on decay kinetics were much less pronounced. The effects of EtOH on IA decay are cell specific among identified neurons of the Aplysia nervous system. The actions of BuOH and HxOH did not mimic these interneuronal differences. These data, coupled with data previously reported by us and others, make it unlikely that EtOH exerts its actions on IA via perturbation of a bulk lipid phase within the membrane.

Ethanol-induced reduction of neuronal calcium currents in Aplysia: an examination of possible mechanisms.

1. Experiments were performed to determine the mechanisms by which ethanol (EtOH) decreases the amplitude of voltage-dependent inward currents through calcium channels in Aplysia neurons. Voltage-clamp protocols used conditioning prepulses of varying amplitude, duration, and frequency, to examine the relationship between prior activity of the channel and EtOH action. Calcium and barium were used as charge carriers, allowing dissociation of effects due to inactivation of calcium channels from other perturbations resulting in the impediment of current flow through the open channel. 2. When Ba2+ was the charge carrier and channel activation was unconfounded by inactivation processes, the reduction of ICa produced by EtOH was independent of the voltage, frequency, or duration of conditioning prepulses. 3. When Ca2+ was the charge carrier, ICa was reduced as a function of conditioning prepulses, in three protocols used. EtOH enhanced this reduction, most probably because of its effects on the inactivation of ICa. Consistent with this interpretation, the time constant of decay of ICa was decreased, and recovery from inactivation was retarded by EtOH. 4. EtOH did not reduce ICa by a change in membrane surface potential, at least at low EtOH concentrations. 5. An analysis of the time course of development of ICa reduction by EtOH showed that it developed slowly, over a matter of minutes. 6. Our data indicate that EtOH does not reduce ICa by direct occlusion of the calcium channel. EtOH affects the inactivation of the calcium current, and this may occur by an action on the channel protein.

Inhibitory action of ethanol on luteinizing hormone secretion by rat anterior pituitary cells in culture.

Ethanol (EtOH) inhibits LH secretion in humans and animals. In these studies we examined the gonadotroph as a possible site of action of EtOH by treating cultured pituitary cells with several concentrations of EtOH (200, 400, or 800 mg/100 ml) for 4 days. Cells were incubated for 2 h with LHRH on the last day of the experiment. Whereas basal LH release was unaffected over the 4 days of EtOH treatment, the LH response to LHRH was inhibited by EtOH in a dose-related fashion. Inhibitory responses at the lowest concentration of EtOH (200 mg/100 ml) occurred inconsistently. At the highest concentration (800 mg/100 ml), the curve of log LHRH dose vs. LH release had a lower maximum and was shifted to the right of that for untreated cells. Total LH content and total number of cells attached to the culture wells were unchanged after EtOH treatment. These data suggest that EtOH at high concentrations can lower the responsiveness of the gonadotroph to LHRH.

Ethanol effects on voltage-dependent membrane conductances: comparative sensitivity of channel populations in Aplysia neurons.

The study of ethanol (EtOH) action is interesting because of its clinical relevance and for the insights it provides into structure-function relationships of excitable membranes. This paper describes the concentration dependencies of various parameters of four currents in Aplysia cells. ICa is the most sensitive of the currents studied. There was a significant reduction of ICa at concentrations of 50 mM EtOH. At low concentrations, the reduction of amplitude was the primary effect of ethanol, with the kinetics and voltage dependency of activation not affected. INa and IA were also affected, but at EtOH levels higher than those which altered ICa. The primary effect of EtOH on INa was a reduction in its amplitude, although the time to peak current flow was increased by EtOH. The effects of EtOH on IA were cell specific and, for the purposes of this paper, we examined the giant metacerebral cell (MCC). In MCC, the primary effect of EtOH on IA was an increase in the time course of inactivation. The time to peak IA was also increased by high concentrations of EtOH, but its amplitude was unaffected even at high concentrations. The delayed rectifier current, IK, was the most EtOH resistant of the currents examined. High EtOH concentrations augmented the amplitude of IK, although even at 600 mM concentrations, the percentage change was only 30%. Our results indicate that the calcium channel is very susceptible to the influence of ethanol and is a serious candidate to be the primary target of EtOH action in the nervous system. The differential sensitivity of voltage-dependent currents and individual components of a given current suggests further experiments to probe the relationship between membrane structure and channel function in excitable membranes.

Antagonism of ethanol-evoked responses by amantadine: A possible clinical application

The behavioral and biochemical effects of amantadine hydrochloride (ATD) on some ethanol (ETOH) mediated responses in rats and mice were studied. Administration of ATD, 0.5 mM/kg, IP, prior to a narcotic dose of ETOH significantly decreased the central depressant action of ETOH, as measured by the duration of ETOH-produced narcosis in mice. The time required for the onset of ETOH-narcosis was significantly delayed in ATD-treated mice compared to controls. Analyses of whole blood and brain ETOH concentrations showed that ATD-treatment prior to ETOH significantly reduced brain content of ETOH from saline-pretreated mice at the time of onset of ETOH narcosis as well as 30 min after ETOH injection without concomitant change in blood ETOH concentrations at the respective time intervals. Administration of ATD, 0.5 mM/kg, IP, reduced voluntary intake of ETOH by rats voluntarily selecting 5% ETOH solution over water as the drinking fluid. There were no changes in cytoplasmic rat liver alcohol dehydrogenase (L-ADH) and mitochondrial aldehyde dehydrogenase (L-ALDH) activities in rats maintained on water or 5% ETOH as the drinking fluid and administered ATD, 0.5 mM/kg, IP, once or identical dose once daily for six consecutive days. However, ATD produced in vitro non-competitive inhibition of both L-ADH and L-ALDH at a concentration range between 10−3 M and 3×10−3 M assay mixture. The results indicate the potency of ATD in negating ETOH-mediated responses measured and suggest for a possible clinical trial for ATD in the management of alcoholic patients provided it is devoid of disulfiram-like reaction in man.

Alkali metal ions and ethanol narcosis in mice.

The effect of an equimolar dose of LiCl, RbCl or CsCl on the depressant action of ethanol (ETOH) was studied as a function of duration of ETOH-mediated narcosis in mice. The alkali metal ions were administered acutely or semi-chronically prior to the administration of a narcotic dose of ETOH. Semi-chronic treatment with RbCl or CsCl resulted in decreased duration of ETOH-mediated narcosis, which was persistent for 5 days after discontinued administration of RbCl and CsCl. Conversely, a prolongation of ETOH-produced narcosis in Li-treated mice occured only when ETOH was shortly administered after sub-chronic regiments of LiCl. Semi-chronic treatment with an equimolar dose of LiCl, RbCl or CsCl produced little changes in the specific activities of liver alcohol and aldehyde dehydrogenase. It is concluded that both Rb+ and Cs+ possess antidepressant properties and might be useful agents in negating the depressant action of ETOH and/or some of its toxic manifestations.