The promiscuous role of the epsilon subunit in GABA(A) receptor biogenesis

The formation of alpha1beta2gamma2epsilon receptors suggests that the epsilon subunit does not displace the single gamma2 subunit in alpha1beta2gamma2 receptors. Thus, epsilon must replace alpha and/or beta subunit(s) if the pentameric receptor structure is to be preserved. To assess the potential for which subunit is replaced in alphabetaepsilon and alphabetagammaepsilon receptors we analyzed the assembly and functional expression of the epsilon subunit with respect to alpha1, beta2 and gamma2 subunits. Using concatenated subunits, we have determined that epsilon is capable of substituting for either (but not both) of the alpha subunits, one of the beta subunits, and possibly the gamma2 subunit. However, the most likely sites at which the epsilon subunit may contribute to receptor function appears to be at position 1 (replaces alpha1) in alphabetagammaepsilon (varepsilon-beta2-alpha1-beta2-gamma2) receptors, or at position 4 (replaces beta2) in alphabetaepsilon (alpha1-beta2-alpha1-varepsilon-beta2) receptors. In both cases, it appears that only a single GABA binding site is present.

Replacement of histidine in position 105 in the alpha(5) subunit by cysteine stimulates zolpidem sensitivity of alpha(5)beta(2)gamma(2) GABA(A) receptors

Zolpidem is a positive allosteric modulator of GABA(A) receptors with sensitivity to subunit composition. While it acts with high affinity and efficacy at GABA(A) receptors containing the alpha(1) subunit, it has a lower affinity to GABA(A) receptors containing alpha(2), alpha(3), or alpha(5) subunits and has a very weak efficacy at receptors containing the alpha(5) subunit. Here, we show that replacing histidine in position 105 in the alpha(5) subunit by cysteine strongly stimulates the effect of zolpidem in receptors containing the alpha(5) subunit. The side chain volume of the amino acid residue in this position does not correlate with the modulation by zolpidem. Interestingly, serine is not able to promote the potentiation by zolpidem. The homologous residues to alpha(5)H105 in alpha(1), alpha(2), and alpha(3) are well-known determinants of the action of classical benzodiazepines. Other studies have shown that replacement of these histidines alpha(1)H101, alpha(2)H101, and alpha(3)H126 by arginine, as naturally present in alpha(4) and alpha(6), leads to benzodiazepine insensitivity of these receptors. Thus, the nature of the amino acid residue in this position is not only crucial for the action of classical benzodiazepines but in alpha(5) containing receptors also for the action of zolpidem.

Differential effect of side of temporal lobe epilepsy on lateralization of hippocampal, temporolateral, and inferior frontal activation patterns during a verbal episodic memory task

The encoding of verbal stimuli elicits left-lateralized activation patterns within the medial temporal lobes in healthy adults. In our study, patients with left- and right-sided temporal lobe epilepsy (LTLE, RTLE) were investigated during the encoding and retrieval of word-pair associates using functional magnetic resonance imaging. Functional asymmetry of activation patterns in hippocampal, inferior frontal, and temporolateral neocortical areas associated with language functions was analyzed. Hippocampal activation patterns in patients with LTLE were more right-lateralized than those in patients with RTLE (P<0.05). There were no group differences with respect to lateralization in frontal or temporolateral regions of interest (ROIs). For both groups, frontal cortical activation patterns were significantly more left-lateralized than hippocampal patterns (P<0.05). For patients with LTLE, there was a strong trend toward a difference in functional asymmetry between the temporolateral and hippocampal ROIs (P=0.059). A graded effect of epileptic activity on laterality of the different regional activation patterns is discussed.

Hippocampal functional connectivity reflects verbal episodic memory network integrity

Using functional magnetic resonance imaging during a verbal memory task, we investigated correlations of signal fluctuations within the hippocampus and ipsilateral frontal as well as temporal areas in temporal lobe epilepsy patients. Declarative memory abilities were additionally examined before and after temporal lobe epilepsy surgery. A significant difference exists in functional connectivity between patients whose mnemonic functions deteriorated and those who remained stable or improved. Univariate analyses showed significantly higher preoperative coupling between the hippocampus and Brodmann area 22 for the group that decreased in verbal learning. We suggest greater coupling to reflect higher functional network integrity. Postoperatively reduced learning ability in patients with higher preoperative coupling underlines the importance of hippocampal interaction with cortical areas for successful memory formation.

BOLD correlates of continuously fluctuating epileptic activity isolated by independent component analysis

Combined EEG/fMRI recordings offer a promising opportunity to detect brain areas with altered BOLD signal during interictal epileptic discharges (IEDs). These areas are likely to represent the irritative zone, which is itself a reflection of the epileptogenic zone. This paper reports on the imaging findings using independent component analysis (ICA) to continuously quantify epileptiform activity in simultaneously acquired EEG and fMRI. Using ICA derived factors coding for the epileptic activity takes into account that epileptic activity is continuously fluctuating with each spike differing in amplitude, duration and maybe topography, including subthreshold epileptic activity besides clear IEDs and may thus increase the sensitivity and statistical power of combined EEG/fMRI in epilepsy. Twenty patients with different types of focal and generalized epilepsy syndromes were investigated. ICA separated epileptiform activity from normal physiological brain activity and artifacts. In 16/20 patients, BOLD correlates of epileptic activity matched the EEG sources, the clinical semiology, and, if present, the structural lesions. In clinically equivocal cases, the BOLD correlates aided to attribute proper diagnosis of the underlying epilepsy syndrome. Furthermore, in one patient with temporal lobe epilepsy, BOLD correlates of rhythmic delta activity could be employed to delineate the affected hippocampus. Compared to BOLD correlates of manually identified IEDs, the sensitivity was improved from 50% (10/20) to 80%. The ICA EEG/fMRI approach is a safe, non-invasive and easily applicable technique, which can be used to identify regions with altered hemodynamic effects related to IEDs as well as intermittent rhythmic discharges in different types of epilepsy.

Concurrent activation of dopamine D1 and D2 receptors is required to evoke neural and behavioral phenotypes of cocaine sensitization

Repeated exposure to psychomotor stimulants produces a striking behavioral syndrome involving repetitive, stereotypic behaviors that occur if an additional exposure to the stimulant is experienced. The same stimulant exposure produces specific alterations in gene expression patterns in the striatum. To identify the dopamine receptor subtypes required for the parallel expression of these acquired neural and behavioral responses, we treated rats with different D1-class and D2-class dopamine receptor agonists and compared the responses of drug-naive rats with those of rats given previous intermittent treatment with cocaine. In rats exposed to repeated cocaine treatment, the effects of a subsequent challenge treatment with either a D1-class agonist (SKF 81297) or a D2-class agonist (quinpirole) were not significantly different from those observed in drug-naive animals: the drugs administered singly did not induce robust stereotyped motor behaviors nor produce significantly striosome-predominant expression of early genes in the striatum. In contrast, challenge treatment with the D1-class and D2-class agonists in combination led to marked and correlated increases in stereotypy and striosome-predominant gene expression in the striatum. Thus, immediately after repeated psychomotor stimulant exposure, only the concurrent activation of D1 and D2 receptor subclasses evoked expression of the neural and behavioral phenotypes acquired through repeated cocaine exposure. These findings suggest that D1-D2 dopamine receptor synergisms underlie the coordinate expression of both network-level changes in basal ganglia activation patterns and the repetitive and stereotypic motor response patterns characteristic of psychomotor stimulant sensitization.

Dopamine inhibition and the adaptation of behavior to ongoing reality

Spontaneous confabulation is a rare memory disorder resulting from orbitofrontal damage or disconnection. Patients act on the basis of memories that do not pertain to the current situation, and are disoriented. No medical treatment is known. Recent studies suggest that subcortical dopaminergic structures are involved in the selection of currently relevant memories. We present a patient who regained the ability to adapt thought and behavior to ongoing reality when treated with risperidone, a dopamine antagonist.

Adenosine A2A receptor gene expression in the normal striatum and after 6-OH-dopamine lesion

Adenosine A2A receptors are present on enkephalinergic medium sized striatal neurons in the rat and have an important function in the modulation of striatal output. In order to establish more accurately whether adenosine transmission is a generalized phenomenon in mammalian striatum we compared the A2A R expression in the mouse, rat, cat and human striatum. Secondly we compared the modulation of enkephalin gene expression and A2A receptor gene expression in rat striatal neurons after 6-OH-dopamine lesion of the substantia nigra. Hybridization histochemistry was performed with a 35S-labelled radioactive oligonucleotide probe. The results showed high expression of A2A adenosine receptor genes only in the medium-sized cells of the striatum in all examined species. In the rat striatum, expression of A2A receptors was not significantly altered after lesion of the dopaminergic pathways with 6-OH-dopamine even though enkephalin gene expression was up-regulated. The absence of a change in A2A receptor gene expression after 6-OH-dopamine treatment speaks against a dependency on dopaminergic innervation. The maintained inhibitory function of A2A R on motor activity in spite of dopamine depletion could be partly responsible for the depression of locomotor activity observed in basal ganglia disorders such as Parkinson's disease.

Comparison of voxel-based 3-D MRI analysis and subtraction ictal SPECT coregistered to MRI in focal epilepsy

While voxel-based 3-D MRI analysis methods as well as assessment of subtracted ictal versus interictal perfusion studies (SISCOM) have proven their potential in the detection of lesions in focal epilepsy, a combined approach has not yet been reported. The present study investigates if individual automated voxel-based 3-D MRI analyses combined with SISCOM studies contribute to an enhanced detection of mesiotemporal epileptogenic foci. Seven consecutive patients with refractory complex partial epilepsy were prospectively evaluated by SISCOM and voxel-based 3-D MRI analysis. The functional perfusion maps and voxel-based statistical maps were coregistered in 3-D space. In five patients with temporal lobe epilepsy (TLE), the area of ictal hyperperfusion and corresponding structural abnormalities detected by 3-D MRI analysis were identified within the same temporal lobe. In two patients, additional structural and functional abnormalities were detected beyond the mesial temporal lobe. Five patients with TLE underwent epileptic surgery with favourable postoperative outcome (Engel class Ia and Ib) after 3-5 years of follow-up, while two patients remained on conservative treatment. In summary, multimodal assessment of structural abnormalities by voxel-based analysis and SISCOM may contribute to advanced observer-independent preoperative assessment of seizure origin.

Relative positioning of diazepam in the benzodiazepine-binding-pocket of GABA receptors

GABA(A) receptors are the major inhibitory neurotransmitter receptors in the brain. Some of them are targets of benzodiazepines that are widely used in clinical practice for their sedative/hypnotic, anxiolytic, muscle relaxant and anticonvulsant effects. In order to rationally separate these different drug actions, we need to understand the interaction of such compounds with the benzodiazepine-binding pocket. With this aim, we mutated residues located in the benzodiazepine-binding site individually to cysteine. These mutated receptors were combined with benzodiazepine site ligands carrying a cysteine reactive group in a defined position. Proximal apposition of reaction partners will lead to a covalent reaction. We describe here such proximity-accelerated chemical coupling reactions of alpha(1)S205C and alpha(1)T206C with a diazepam derivative modified at the C-3 position with a reactive isothiocyanate group (-NCS). We also provide new data that identify alpha(1)H101C and alpha(1)N102C as exclusive sites of the reaction of a diazepam derivative where the -Cl atom is replaced by a -NCS group. Based on these observations we propose a relative positioning of diazepam within the benzodiazepine-binding site of alpha(1)beta(2)gamma(2) receptors.