Udo Bonnet

Lack of efficacy of naltrexone in the prevention of alcohol relapse: results from a German multicenter study.

In a placebo-controlled, double-blind German multicenter study (seven sites) the efficacy of naltrexone as an adjunctive treatment in alcoholism to maintain abstinence was assessed for 12 weeks. A total of 171 detoxified patients (97.7% met the DSM-III-R criteria for alcohol dependence) were included. Patients had been abstinent for a mean of 19.5 ± 9.4 days at study entry. Eighty-four and 87 patients were randomized to receive naltrexone (50 mg/day) and placebo, respectively. Each site was instructed to provide its usual psychosocial alcohol treatment program. The primary effectiveness measure was the time to first heavy drinking as derived from self-reports of drinking (timeline-follow-back method). Secondary effectiveness measures included time to first drink, amount of alcohol consumption, intensity of craving, severity of alcoholism problems, and liver enzymes. Thirty-three (38%) placebo patients and 28 (33%) naltrexone patients discontinued the study. At endpoint, 62% of the patients in each group did not have an episode of heavy drinking. Also, there were no significant differences between the study groups concerning secondary effectiveness measures as well as compliance and adverse clinical events—with the exception of the γ-GT, which was significantly greater reduced in the naltrexone group throughout the study. Based upon an intention-to-treat population, this study confirms the safety but not the efficacy of naltrexone in prevention of alcohol relapse. Nevertheless, the question arises whether self-reports of drinking are more reliable than γ-GT as a measure of recent alcohol consumption.

Intracellular pH modulates spontaneous and epileptiform bioelectric activity of hippocampal CA3-neurones

A growing body of evidence hints at intracellular free protons to be involved in the modulation of electric activity of cortical neurones. In this study we demonstrate that application of the weak acid propionate (2.5-20 mM) transiently lowers intracellular pH (pH(i)) of BCECF-AM loaded CA3-neurones in hippocampal slices. The predictability of this acidification prompted us to use propionate as a tool to investigate effects of pH(i) on spontaneous bioelectric activity (SBA) and epileptiform activity (EA, induced by bicuculline, caffeine or low magnesium) of CA3 neurones: SBA and EA were transiently suppressed by 2-20 mM propionate - coinciding with the transient neuronal acidification. As activation of Na(+)/H(+)-exchangers (NHE) is involved in the recovery from neuronal acidosis and NHE-inhibition alone is known to increase the activity of intracellular free protons of hippocampal neurones, we tested the effect of the NHE-blockers amiloride (0.5-1 mM) or HOE642 (200 microM) on SBA and EA of CA3-neurones. Long-term application of NHE-inhibitors alone continuously suppressed SBA and EA, which recovered during additional exposure to the weak base trimethylamine (5-10 mM). Simultaneous administration of propionate and NHE-blockers intensified the inhibition of neuronal activity. Together, these results indicate that intracellular acidification inhibits bioelectric activity of hippocampal CA3-neurones. This supports the hypothesis that pH(i) contributes to the control of cortical excitability.

Selective inhibition of the Na+/H+ exchanger type 3 activates CO2/H+-sensitive medullary neurones.

Abstract Hypercapnia as well as lowered intracellular pH (pHi) increase the bioelectric activity of CO2/H+-sensitive neurones (VLNcs) of the ventrolateral medulla oblongata. Here we describe that immunoreactive Na+/H+ exchanger (NHE3) is present in ventrolateral neurones from medullary organotypic cultures (obex level). To test whether VLNcs can be acidified and thereby activated by inhibition of NHE3, we used the novel high-affinity NHE3-inhibitors S1611 and S3226. Both drugs raised the firing rates of VLNcs to at least 150% of the control values, and depolarized membrane potential by up to 15 mV at concentrations (0.5–1 µmol/l) suitable for selective inhibition of NHE3. The changes in bioelectric activity strongly resembled the responses to hypercapnia (PCO2: 60–100 mmHg). In BCECF-AM-loaded cultures a subfraction of ventrolateral VLNcs was found to be intracellularly acidified by 0.05–0.1 pH units following treatment with S1611; the time course of this acidification was similar to that evoked by hypercapnia. All drug effects were sustained and readily reversible upon washing. Non-CO2/H+-responsive medullary neurones as well as hippocampal CA3 neurones were unaffected by up to 20 µmol/l S1611. It is concluded that the selective inhibition of NHE3 acidifies and activates CO2/H+-sensitive neurones within the ventrolateral medulla oblongata.

Treatment of acute alcohol withdrawal with gabapentin: results from a controlled two-center trial.

A few case reports and data from animal experiments point to a possible efficacy of gabapentin (GP) in the treatment of alcohol withdrawal syndrome (AWS). Because of ethical considerations, the efficacy of GP in acute AWS was tested in an add-on fashion to clomethiazole (CLO). Given that the symptom-triggered amount of CLO required to limit AWS within the first 24 hours is related to the severity of AWS, we tested this amount of CLO during placebo (P) or GP (400 mg qid) under double blind, randomized conditions. Sixty-one patients (P = 29/GP = 32) suffering from alcohol dependence (ICD-10) and without any other psychiatric condition or psychotropic medication were included. The groups were not significantly different in baseline characteristics (eg, demographic data, severity of AWS). Both ITT and completer analyses revealed no significant differences between the groups considering the primary effectiveness measure: amount of CLO required in the first 24 hours (P = 6.1 ± 5.4/GP = 6.2 ± 4.7 capsules). In addition, premature discontinuations (P = 3/GP = 2) and decreases in Mainz Alcohol Withdrawal Scores were not significantly different in the first 48 hours of AWS (secondary effectiveness measures). Tolerability of combined CLO/GP was studied throughout the whole treatment comprising a 5-day lasting reduction part subsequent to the first 48 hours. Throughout the whole 7-day treatment a total of 5 and 2 patients dropped out and 6 and 5 patients reported adverse clinical events in the P and GP groups, respectively. All together, GP (400 mg qid) was no better than P in saving initial consumption of CLO or decreasing initial Mainz Alcohol Withdrawal Scores suggesting that GP was ineffective in the management of acute AWS in this model. The combination of GP and CLO was safe.

Carbonic anhydrase inhibitor sulthiame reduces intracellular pH and epileptiform activity of hippocampal CA3 neurons.

Summary:  Purpose: Sulthiame is a carbonic anhydrase (CA) inhibitor with an anticonvulsant effect in the treatment of benign and symptomatic focal epilepsy in children. The aim of the study was to elucidate the mode of action of sulthiame with respect to possible changes of intracellular pH (pHi) that might develop along with sulthiames anticonvulsant properties.

Modulation of voltage-gated channel currents by harmaline and harmane

1 Harmala alkaloids are endogenous substances, which are involved in neurodegenerative disorders such as M. Parkinson, but some of them also have neuroprotective effects in the nervous system. 2 While several sites of action at the cellular level (e.g. benzodiazepine receptors, 5‐HT and GABAA receptors) have been identified, there is no report on how harmala alkaloids interact with voltage‐gated membrane channels. 3 The aim of this study was to investigate the effects of harmaline and harmane on voltage‐activated calcium‐ (ICa(V)), sodium‐ (INa(V)) and potassium (IK(V))‐channel currents, using the whole‐cell patch‐clamp method with cultured dorsal root ganglion neurones of 3‐week‐old rats. Currents were elicited by voltage steps from the holding potential to different command potentials. 4 Harmaline and harmane reduced ICa(V), INa(V) and IK(V) concentration‐dependent (10–500 μM) over the voltage range tested. ICa(V) was reduced with an IC50 of 100.6 μM for harmaline and by a significantly lower concentration of 75.8 μM (P<0.001, t‐test) for harmane. The Hill coefficient was close to 1. Threshold concentration was around 10 μM for both substances. 5 The steady state of inhibition of ICa(V) by harmaline or harmane was reached within several minutes. The action was not use dependent and at least partly reversible. 6 It was mainly due to a reduction in the sustained calcium channel current (ICa(L+N)), while the transient voltage‐gated calcium channel current (ICa(T)) was only partially affected. 7 We conclude that harmaline and harmane are modulators of ICa(V) in vitro. This might be related to their neuroprotective effects.

CO2/HCO3-withdrawal from the bath medium of hippocampal slices: biphasic effect on intracellular pH and bioelectric activity of CA3-neurons

Many studies analyzing interactions of pH and bioelectric activity focus on changes of the extracellular pH, whereas data concerning central neuronal excitability and intracellular pH (pHi) are rare. Here, we report on the spontaneous bioelectric activity and epileptiform activity of CA3-neurons during a procedure which changed pHi. As monitored in BCECF-AM loaded cells, the change from a CO2/HCO3(-)-buffered to a HEPES-buffered medium (CO2/HCO3(-)-withdrawal, hereafter termed W) was associated with a transient intracellular alkalosis (delta pH = 0.2 +/- 0.04) which preceded a sustained intracellular acidosis (delta pH = 0.4 +/- 0.04). Coinciding with this W-induced biphasic shift of pHi a biphasic alteration of spontaneous bioelectric activity was recorded: as a rule, an up to 30 min lasting increase (excitatory phase) preceded a typical sustained suppression (inhibitory phase). This biphasic action was also observed using various in vitro-epilepsy-models (bicuculline, penicillin, caffeine): epileptiform discharges were completely suppressed after an initial increase in frequency. This modulation of bioelectric activity was unlikely due to alterations of the postsynaptic GABA-system as hyperpolarizing GABAA- and GABAB-responses of CA3-neurons were hardly affected. In the majority of the neurons, the initial increase of spontaneous bioelectric activity (excitatory phase) culminated in transient burst periods lasting 5-30 min. These transient burst periods were blocked by NMDA- or AMPA-antagonists: DL-2-amino-5-phosphonovalerate (APV, 50 microM) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 50 microM). The calcium-antagonist verapamil (50 microM) reduced amplitudes of depolarizations and duration of the transient burst periods. The results suggest that the biphasic alteration of pHi modulates the susceptibility of glutamate receptors and voltage-gated calcium-channels, which leads to respective changes of bioelectric activity.

CO2-sensitive medullary neurons : activation by intracellular acidification

BIOELECTRIC activity of CO2-sensitive, ventrolateral medullary neurons (VLNCS) in organotypic cultures from the obex level of newborn rats was tested during changes of the intracellular pH (pHi) measured in BCECF-AM loaded cultures. Hypercapnia (pCO2 80–100 mmHg) reduced pHi by 0.15 ± 0.06 units and stimulated neuronal discharges. Replacement of CO2/HCO3− in the bath by HEPES (26 mM, pH 7.4) for 10 min acidified pHi (0.07 ± 0.03 units) and also excited VLNCS. Ammonium chloride (10 mM, 1 min) initially alkalized (0.1 ± 0.04) and thereafter acidified pHi (0.06 ± 0.03), while the extracellular pH was first acidified and then alkalized. This resulted in neuronal discharge which were first suppressed and then accelerated. The findings strongly suggest that intracellular rather than extra-cellular acidification activates CO2-sensitive neurons.

Moclobemide reduces intracellular pH and neuronal activity of CA3 neurones in guinea-pig hippocampal slices—implication for its neuroprotective properties

Mechanisms underlying the neuroprotective properties of the weak MAO-A inhibitor moclobemide are not understood. Increasing evidence suggests that a moderate increase in intracellular free protons may contribute to neuroprotective properties due to a proton-mediated decrease in neuronal activity. Therefore, we studied effects of 10-700 microM moclobemide (i) on the intracellular pH (pH(i)) of BCECF-AM loaded CA3 neurones as well as (ii) on spontaneous action potentials and epileptiform activity (induced by bicuculline-methiodide, caffeine, or 4-aminopyridine) of CA3 neurones in the stratum pyramidale. Moclobemide-concentrations of > or = 300 microM reversibly reduced the steady-state pH(i) by up to 0. 25 pH-units within 5-20 min. Simultaneously, the frequency of spontaneous action potentials and epileptiform discharges became depressed. Moclobemide also abolished 4-aminopyridine-induced GABA-mediated hyperpolarisations suggesting that the inhibitory and acidifying effects of moclobemide do not result from an amplification of the GABA system. The stronger MAO-A inhibitors clorgyline or pargyline (both 10 microM) mimicked the moclobemide-effects. Investigating effects on pH(i)-regulation we found that 700 microM moclobemide impaired the recovery from intracellular acidification elicited by an ammonium prepulse which demonstrates an impairment of transmembrane acid extrusion. We suggest that the latter effect is responsible for the moderate decrease in the steady-state pH(i) which in turn reduced neuronal activity. This mechanism may substantially contribute to the neuroprotective properties of moclobemide.

Vermal atrophy of alcoholics correlate with serum thiamine levels but not with dentate iron concentrations as estimated by MRI

AbstractChronic alcohol consumption is frequently accompanied by cerebellar degeneration. The exact aetiology of alcoholic cerebellar degeneration is still a matter of debate. The aim of the present study was to investigate whether patients with chronic alcohol consumption exhibit a decrease in dentate nuclei intensity as measured by MRI, and if so, whether this decrease correlates with cerebellar atrophy as revealed by MR imaging or with clinical signs of cerebellar ataxia. A decrease in dentate nuclei intensity would indirectly indicate that iron accumulation, and therefore, oxidative stress may play a role in alcoholic cerebellar degeneration.MRI of 45 alcoholics and 44 ageand sex–matched healthy control subjects was performed using a 3D–T1–weighted fast low angle shot (FLASH) echo sequence. Signal intensities of the dentate nuclei and cerebellar white matter were bilaterally measured. Planimetric measurements of cerebellar size were performed using a 3D–T1–weighted magnetization prepared rapid acquisition gradient echo (MPRAGE) sequence.Results demonstrated that dentate nuclei intensity was not significantly decreased in patients with chronic alcohol consumption (mean ± SD signal intensity 65.36 ± 13.0) if compared with control subjects (mean ± SD signal intensity 68.95 ± 9.4) (p = 0.15). Dentate nuclei intensity did not correlate with cerebellar size neither in control subjects nor in alcoholics. In contrast, vitamin B1 level correlated with cerebellar size in alcoholics even if the vitamin B1 concentration was within normal values (r = 0.344, p = 0.028). These results support the view that thiamine deficiency rather than direct neurotoxic effects of alcohol is the main causative factor for the development of alcoholic cerebellar degeneration.