The GHB analogue n-(4-trifluoromethyl)-4-methoxybutanamide prevents the behavioral and neurodegenerative consequences of repeated episodes of alcohol withdrawal, in rats.

Abrupt cessation of chronic consumption of alcohol produces a severe withdrawal syndrome, characterized by behavioral signs (increased anxiety, tremor, hyperactivity and seizures, conspicuous memory deficits) associated with neurodegeneration, particularly in the hippocampus and in the hippocampal cholinergic fiber network. The mechanism appears to be a classical excitotoxic cascade, with overactivation of the glutamatergic/NMDA neurotransmission associated with concurrently diminished GABAergic neurotransmission. Gamma-hydroxybutyric acid (GHB) functions as a neurotransmitter in the CNS, with specific high affinity binding sites in defined brain structures, including the hippocampus, and with affinity also for GABA B receptors, alike present in the hippocampus. GHB has long been used for the treatment of alcohol dependence, where it effectively decreases ethanol craving and consumption as well as withdrawal symptoms, likely by a substitution mechanism. Moreover, it exerts a protective effect against brain damage produced by traumatic, excitotoxic or ischemic injuries. N-(4-trifluoromethylbenzyl)-4-methoxybutanamide (GET73) is a GHB analogue with more potent and longer-lasting activity than the parent compound. Aim of the present research was to study the possible protective effect of GET73 against the consequences of repeated episodes of alcohol withdrawal in chronically alcohol-consuming rats. METHODS and TREATMENT. After 1 week of adaptation to our housing conditions, adult male rats of a Wistar Kyoto strain had tap water substituted with an alcohol aqueous solution (6% by volume) as ad libitum beverage. Control rats received tap water containing sucrose of same caloric content. At the end of the 3rd, 4th and 5th week, alcohol solution -and sucrose solution in controls- were withdrawn for 24 hr and replaced with tap water. At the end of the last withdrawal period, rats were observed for gross behavior and open-field behavior. After the behavioral observation, rats were killed under ether anesthesia, and brains were removed for histological examination. Alcohol-consuming rats were randomly assigned to i.p. treatment either with GET73 (1, 5 or 10 mgkg-1, at the beginning of each withdrawal episode, and again 12 hr later and 30 min before behavioral testing) or with the vehicle. The research protocol had been approved by the Animal Experimentation Ethical Committee of the University of Modena and Reggio Emilia. RESULTS. At the end of the 3rd alcohol-withdrawal episode, vehicle-treated rats were excited, hyperactive, squeaking, aggressive and attemping to bite when handled; in the open-field, grooming and ambulation were increased, compared with non-alcohol-consuming rats. GET73 produced a reduction of ambulation (inner ond outer crossing), rearing and grooming (vehicle-treated: inner cross. 14.8 +/- 6.5; outer cross. 53.0 +/- 25.7; rearings 7.8 +/- 4.2; grooming 10.5 +/-5.2 (m +/- S.D.). GET73 10 mg/kg: inner cross. 3.6 +/- 2.3 (F = 16.77; P = 0.000); outer cross. 13.4 +/-11.0 (F = 9.26; P = 0.000); rearings 3.8 +/- 2.2 (F = 11.1; P= 0.000); grooming 0.0 (F= 16.80; P= 0.000)(ANOVA followed by S.N.K. test)). The histological picture in abstinent rats treated with the vehicle was characterized by a severe damage of the hippocampal CA1 subfield, with almost complete absence of vital neurons, demyelinization, marked reduction of the number of synapses, and astrogliosis. The treatment with GET73 produced a dose-dependent protection: with the highest dose (10 mg/kg) the number of vital neurons and synapses was minimally reduced and astrogliosis was quite limited. CONCLUSIONS. These results show that in chronically alcohol-consuming rats, the behavioral and neurodegenerative consequences of repeated episodes of alcohol withdrawal are prevented by the GHB analogue N-(4-trifluoromethyl)-4-methoxybutanamide (GET73). The possible mechanisms may likely involve (i) reduced release of excitatory amino acids as a consequence of the activation of both GHB and GABAB receptors, (ii) hyperpolarisation of hippocampal neurons by activating GABAB receptors and by increasing K+ conductance.