Involvement of the limbic basal ganglia in ethanol withdrawal convulsivity in mice is influenced by a chromosome 4 locus.

Physiological dependence and associated withdrawal episodes are thought to constitute a motivational force that sustains ethanol (alcohol) use/abuse and may contribute to relapse in alcoholics. Although no animal model duplicates alcoholism, models for specific factors, like the withdrawal syndrome, are useful for identifying potential genetic and neural determinants of liability in humans. We generated congenic mice that confirm a quantitative trait locus (QTL) on chromosome 4 with a large effect on predisposition to alcohol withdrawal. Using c-Fos expression as a high-resolution marker of neuronal activation, congenic mice demonstrated significantly less neuronal activity associated with ethanol withdrawal than background strain mice in the substantia nigra pars reticulata (SNr), subthalamic nucleus (STN), rostromedial lateral globus pallidus, and ventral pallidum. Notably, neuronal activation in subregions of the basal ganglia associated with limbic function was more intense than in subregions associated with sensorimotor function. Bilateral lesions of caudolateral SNr attenuated withdrawal severity after acute and repeated ethanol exposures, whereas rostrolateral SNr and STN lesions did not reduce ethanol withdrawal severity. Caudolateral SNr lesions did not affect pentylenetetrazol-enhanced convulsions. Our results suggest that this QTL impacts ethanol withdrawal via basal ganglia circuitry associated with limbic function and that the caudolateral SNr plays a critical role. These are the first analyses to elucidate circuitry by which a confirmed addiction-relevant QTL influences behavior. This mouse QTL is syntenic with human chromosome 9p. Given the growing body of evidence that a gene(s) on chromosome 9p influences alcoholism, our results can facilitate human research on alcohol dependence and withdrawal.

Activity of hippocampal adult-born neurons regulates alcohol withdrawal seizures.

Alcohol withdrawal (AW) after chronic alcohol exposure produces a series of symptoms, with AW-associated seizures being among the most serious and dangerous. However, the mechanism underlying AW seizures has yet to be established. In our mouse model, a sudden AW produced 2 waves of seizures: the first wave includes a surge of multiple seizures that occurs within hours to days of AW, and the second wave consists of sustained expression of epileptiform spikes and wave discharges (SWDs) during a protracted period of abstinence. We revealed that the structural and functional adaptations in newborn dentate granule cells (DGCs) in the hippocampus underlie the second wave of seizures but not the first wave. While the general morphology of newborn DGCs remained unchanged, AW increased the dendritic spine density of newborn DGCs, suggesting that AW induced synaptic connectivity of newborn DGCs with excitatory afferent neurons and enhanced excitability of newborn DGCs. Indeed, specific activation and suppression of newborn DGCs by the chemogenetic DREADD method increased and decreased the expression of epileptiform SWDs, respectively, during abstinence. Thus, our study unveiled that the pathological plasticity of hippocampal newborn DGCs underlies AW seizures during a protracted period of abstinence, providing critical insight into hippocampal neural circuits as a foundation to understand and treat AW seizures.

The effects of chronic ethanol administration on amygdala neuronal firing and ethanol withdrawal seizures.

Physical dependence on ethanol results in an ethanol withdrawal (ETX) syndrome including susceptibility to audiogenic seizures (AGS) in rodents after abrupt cessation of ethanol. Chronic ethanol administration and ETX induce functional changes of neurons in several brain regions, including the amygdala. Amygdala neurons are requisite elements of the neuronal network subserving AGS propagation during ETX induced by a subacute "binge" ethanol administration protocol. However, the effects of chronic ethanol administration on amygdala neuronal firing and ETX seizure behaviors are unknown. In the present study ethanol (5g/kg) was administered intragastrically in Sprague-Dawley rats once daily for 28days [chronic intermittent ethanol (CIE) protocol]. One week later the rats began receiving ethanol intragastrically three times daily for 4days (binge protocol). Microwire electrodes were implanted prior to CIE or on the day after CIE ended to record extracellular action potentials in lateral amygdala (LAMG) neurons. The first dose of ethanol administered in the binge protocol following CIE treatment did not alter LAMG neuronal firing, which contrasts with firing suppression seen previously in the binge protocol alone. These data indicate that CIE induces neuroadaptive changes in the ETX network which reduce LAMG response to ethanol. LAMG neuronal responses to acoustic stimuli prior to AGS were significantly decreased during ETX as compared to those before ethanol treatment. LAMG neurons fired tonically throughout the tonic convulsions during AGS. CIE plus binge treatment resulted in a significantly greater mean seizure duration and a significantly elevated incidence of death than was seen previously with the binge protocol alone, indicating an elevated seizure severity following chronic ethanol administration.

Role of the amygdala in ethanol withdrawal seizures.

Ethanol withdrawal (ETX) after induction of ethanol dependence results in a syndrome that includes enhanced seizure susceptibility. During ETX in rodents, generalized audiogenic seizures (AGS) can be triggered by intense acoustic stimulation. Previous studies have implicated specific brainstem nuclei in the neuronal network that initiates and propagates AGS during ETX. Although ethanol and ETX are known to affect amygdala neurons, involvement of the amygdala in the network subserving AGS is unclear. Since ethanol and ETX affect N-methyl-d-aspartate (NMDA) receptors in the amygdala, the present study evaluated the effect of focally microinjecting a NMDA antagonist into the amygdala of rats treated with a binge protocol (intragastric administration of ethanol 3 times daily for 4 days). Separate experiments examined extracellular neuronal firing in the amygdala. Cannulae or microwire electrodes were chronically implanted into the amygdala, and changes in seizure behaviors and/or extracellular action potentials were evaluated. Bilateral focal microinjection of a NMDA antagonist, 2-amino-7-phosphonoheptanoate (AP7), into either central nucleus or lateral nucleus of the amygdala (LAMG) significantly reduced AGS. The doses of AP7 and time course of effect were similar in each site, suggesting that both amygdala nuclei participate in the AGS network. Acoustic responses of LAMG neurons were significantly decreased 1 h after the first ethanol dose and also during ETX, as compared to pre-binge controls. However, LAMG neurons consistently exhibited rapid tonic firing during the generalized tonic convulsions of AGS. These findings suggest a critical role of the amygdala in the ETX seizure network in generating tonic convulsions during AGS.

The influence of ethanol intake and its withdrawal on the anticonvulsant effect of alpha-tocopherol in the penicillin-induced epileptiform activity in rats.

Previous studies proposed the existence of a relationship between epilepsy and ethanol. Ethanol may have either proconvulsive or anticonvulsive effects on epileptic activity in different experimental epilepsy models. The influence of high dose ethanol intake and its withdrawal on the anticonvulsant effect of alpha-tocopherol was examined after intracortical injection of penicillin (500 units) to induce epileptiform activity. Thirty minutes after penicillin injection, the most effective dose of alpha-tocopherol (500 mg/kg) was administrated intramuscularly (i.m.). Ethanol-treated rats received a daily dose of 9.0 g/kg of 30% ethanol solution via an oesophageal probe for 15 days. All rats in the withdrawal group were anesthetized for induction of penicillin-induced epileptiform activity 28 h after the last ethanol administration. The epileptiform activity was verified by electrocorticographic (ECoG) recordings. Ethanol, in a dose of 9 g/kg, significantly decreased the mean frequency of penicillin-induced epileptiform ECoG activity without changing the amplitude. The mean frequency of ECoG activity was decreased in the 60 and 70 min period from penicillin injection in the ethanol-treated+alpha-tocopherol and ethanol withdrawal+alpha-tocopherol groups compared with the penicillin-injected (500 units, i.c.) group, respectively. alpha-Tocopherol was more effective in decreasing the mean frequency of epileptiform activity in the ethanol+alpha-tocopherol group than in other alpha-tocopherol administrated groups. Ethanol withdrawal caused an increase in frequency of epileptiform activity in the withdrawal+alpha-tocopherol group compared with other alpha-tocopherol administrated groups. alpha-Tocopherol did not affect the amplitude of epileptiform activity in any group. Possible mechanisms of ethanol influence on the neuroprotective actions of alpha-tocopherol are still a crucial issue associated with epilepsy.

Neurons in the periaqueductal gray are critically involved in the neuronal network for audiogenic seizures during ethanol withdrawal.

The periaqueductal gray (PAG) is implicated in the network subserving audiogenic seizures (AGS). AGS are seen during ethanol withdrawal (ETX), and the present study examined effects of focal NMDA receptor blockade in PAG during ETX and PAG neuronal firing changes associated with ETX. Bilateral cannulae or microwire electrodes were chronically implanted into PAG. Ethanol was administered intragastrically at 8-h intervals for 4 days, resulting in AGS susceptibility during ETX. Microinjection of a competitive NMDA receptor antagonist, DL-2-amino-7-phosphonoheptanoic acid (AP7) (2 and 5 but not 1 nmol/side), into the PAG suppressed AGS, in part, reversibly. In microwire experiments spontaneous and acoustically evoked PAG neuronal responses in behaving rats were reduced significantly 1 h after initial administration of ethanol. During ETX, when the animals were susceptible to AGS, significant increases in spontaneous and acoustically evoked PAG neuronal firing occurred. PAG neurons exhibited burst firing 2-4 s prior to the tonic-clonic phase of AGS and tonic repetitive firing during this seizure phase, which ceased during post-ictal depression. Increased NMDA receptor function in PAG may be important to the aberrant PAG neuronal firing in AGS, since previous studies observed upregulation of NMDA receptors during ETX, and the present study observed that focal microinjection of a NMDA antagonist into PAG blocked AGS.

The effects of alcohol intake and withdrawal on the seizures frequency and hippocampal morphology in rats with epilepsy.

The aim of our study, using the pilocarpine model of epilepsy, was to investigate the effects of alcohol administration and withdrawal on the spontaneous recurrent seizures (SRSs). Four groups of adult, male Wistar rats were studied: (A). control rats (n=10), received neither pilocarpine nor alcohol, (B). alcohol-treated rats (n=10), received a daily dose of 3.0 g x kg(-1) of a 30% alcohol solution via an oesophagic probe for 30 days, (C). rats with epilepsy (n=10), (D). rats with epilepsy with alcohol intake (n=10). SRSs were induced by a single dose of pilocarpine (i.p.) and the basal frequency of SRSs was video monitored (24h per day) for 30 days. Following this period, the animals of group D received a daily dose of alcohol solution as described above and at the end of this period, alcohol administration was stopped and the seizure frequency was assessed for more 30 days. The basal seizure frequency observed in groups C and D during the first 30 days was 2.2+/-1.8 seizures per week per animal. In group D, it was observed an increase to 12.2+/-5.8 during the first 2 weeks of alcohol administration. During the last 2 weeks of alcohol administration, the number of SRSs returned to the previous basal level. During alcohol withdrawal the seizure frequency increased to 14.3+/-7.4 seizures per week per animal for the first 2 weeks, and returned to the basal level in the remaining period of observation. The Neo-Timm and Nissl staining of hippocampal formation and of the dentate gyrus in rats with epilepsy showed a cell loss in the hippocampal subfield CA1 and in the hillus of dentate gyrus. In rats with epilepsy with alcohol intake, we observed a cell loss in hippocampal subfields CA3 and hillus of the dentate gyrus, with significant neuronal death in subfield CA1, when compared with control animals. The alcohol withdrawal syndrome is a crucial event for the development of functional and neuropathological alterations associated with epilepsy.

Low sodium excretion in SIADH patients with low diuresis.

UNLABELLED: It is well known that during low diuresis or low effective circulating volume, salt excretion is low. The aim of this study was to find out whether salt excretion, expressed as either urinary sodium concentration (UNa) or fractional sodium excretion (FENa), and the combined use of FENa and fractional urea excretion (FEurea) still differentiate between hyponatremic SIADH and hyponatremic salt depletion (SD) patients when diuresis is low. The relationships between UNa, FENa and diuresis, indirectly estimated by the urinary to plasma creatinine ratio (U/P creat), were studied in 42 hyponatremic SIADH patients, 21 hyponatremic SD patients and 66 normonatremic controls (CO) of similar age and sex ratio. There was no significant relationship between UNa and U/P creat either in SIADH or in SD or CO patients. FENa and U/P creat were inversely correlated, both in CO (r = -0.72; p < 0.001) and in SIADH (r = -0.68; p < 0.001). SIADH and SD patients can be fairly well differentiated from one another using FENa and U/P creat. Even with high U/P creat values, SIADH patients, despite a sharp decrease in their FENa values, presented still higher FENa values than SD patients did (mean FENa = 0.3 +/- 0.2% in SIADH and 0.1 +/- 0.04% in SD; p < 0.05). However, FENa values of SIADH patients with low diuresis (mean FENa = 0.3 +/- 0.2% for a mean U/P creat = 191 +/- 40) are indistinguishable from those of SD patients with normal urine volumes (mean FENa = 0.2 +/- 0.2% for a mean U/P creat = 92 +/- 30). The combined use of FENa and FEurea remains a reliable way to discriminate SD patients and SIADH patients, as far as the differential limit value for FENa is narrowed to a value of 0.15%, for hyponatremic patients with U/P creat >140. CONCLUSION: In SIADH, FENa values are lower than 0.5%, as soon as U/P creat exceeds a value of 180. In SD patients with U/P creat values exceeding 140, FENa is lower than 0.15% and FEurea lower than 45%.