Jacqueline Scuvée-Moreau

Effect of various antidepressant drugs on the spontaneous firing rate of locu coeruleus and dorsal raphe neurons of the rat

The spontaneous firing rate of the noradrenergic neurons of the locus coeruleus and of the serotonergic neurons of the dorsal raphe was recorded with extracellular microelectrodes in chloral hydrate-anesthetized rats. A quantitative comparison of the effect of five tricyclic antidepressants, of tranylcypromine and of mianserin on the spontaneous activity of these two types of cells was performed. All drugs tested, except mianserin reduced the frequency of discharge of the noradrenergic neurons. Intravenous perfusion of the drugs allowed the doses required for inhibition of firing to 50% of the baseline rate (ID50) to be determined. Secondary aminated antidepressants (desipramine and nortriptyline) were more potent inhibitors than their tertiary aminated analogues (imipramine, chlorimipramine and amitriptyline). All drugs tested, except desipramine decreased the rate of firing of the serotonergic cells. In this case, the tertiary aminated antidepressants were much more potent than their secondary analogues. Mianserin was only active at very high doses. These results are in good agreement with the relative potencies of the tricyclic antidepressants for blocking the uptake of noradrenaline and serotonin into central and peripheral neurons.

SK channels control the firing pattern of midbrain dopaminergic neurons in vivo

A vast body of experimental in vitro work and modelling studies suggests that the firing pattern and/or rate of a majority of midbrain dopaminergic neurons may be controlled in part by Ca2+‐activated K+ channels of the SK type. However, due to the lack of suitable tools, in vivo evidence is lacking. We have taken advantage of the development of the water‐soluble, medium potency SK blocker N‐methyl‐laudanosine (CH3‐L) to test this hypothesis in anaesthetized rats. In the lateral ventral tegmental area, CH3‐L iontophoresis onto dopaminergic neurons significantly increased the coefficient of variation of their interspike intervals and the percentage of spikes generated in bursts as compared to the control condition. The effect of CH3‐L persisted in the presence of a specific GABAA antagonist, suggesting a direct effect. It was robust and reversible, and was also observed in the substantia nigra. Control experiments demonstrated that the effect of CH3‐L could be entirely ascribed to its blockade of SK channels. On the other hand, the firing pattern of noradrenergic neurons was much less affected by CH3‐L. We provide here the first demonstration of a major role of SK channels in the control of the switch between tonic and burst firing of dopaminergic neurons in physiological conditions. This study also suggests a new strategy to develop modulators of the dopaminergic (DA) system, which could be of interest in the treatment of Parkinsons disease, and perhaps other diseases in which DA pathways are dysfunctional.

Modulation of Small Conductance Calcium-Activated Potassium (SK) Channels: A New Challenge in Medicinal Chemistry

Small conductance calcium-activated potassium (SK) channels are found in many types of neurons as well as in some other cell types. These channels are selective for K(+) and open when intracellular Ca(2+) rises to omega 500 nM. In neurons, this occurs during and after an action potential. Activation of SK channels hyperpolarizes the membrane, thus reducing cell excitability for several tens or hundreds of milliseconds. This phenomenon is called a afterhyperpolarization (AHP). Three subtypes of SK channels (SK1, SK2, SK3) have been cloned and exhibit a differential localization in the brain. SK channels may play a role in physiological and pathological conditions. They may be involved in the control of memory and cognition. Moreover, they are heavily expressed in the basal ganglia (in particular in the substantia nigra, pars compacta) and in the limbic system, suggesting that they may modulate motricity and emotional behaviour. Based on these facts, SK channel subtypes may be a suitable target for developing novel therapeutic agents, but more work is needed to validate these targets. Hence, there is a great need for selective ligands. Moreover, although the risk of peripheral side-effects for SK channel modulators appears to be low, some questions remain to be investigated. Currently, different molecules are known as SK channel modulators. Apamin is a very potent peptidic agent; it produces a strong blockade of these targets which is only very slowly reversible and it has limited selectivity. Dequalinium was found to be an effective blocker. Different chemical modulations on the dequalinium structure led to the discovery of highly potent bis-quinolinium derivatives such as UCL 1684. Other bis-(2-amino-benzimidazole) derivatives are in development. On the other hand, quaternary salts of bicuculline were reported to be effective in inhibiting AHPs. More recent developments on structurally-related molecules revealed that methyl-laudanosine is a new interesting tool for exploring SK channel pharmacology. Finally, a family of compounds has been shown to facilitate SK channel opening. Such compounds may be useful in treating disorders involving neuronal hyperexcitability.

The KCNQ Channel Opener Retigabine Inhibits the Activity of Mesencephalic Dopaminergic Systems of the Rat

Homo- and heteromeric complexes of KCNQ channel subunits are the molecular correlate of the M-current, a neuron-specific voltage-dependent K+ current with a well established role in control of neural excitability. We investigated the effect of KCNQ channel modulators on the activity of dopaminergic neurons in vitro and in vivo in the rat ventral mesencephalon. The firing of dopaminergic neurons recorded in mesencephalic slices was robustly inhibited in a concentration-dependent manner by the KCNQ channel opener N-(2-amino-4-(4-fluorobenzylamino)-phenyl) carbamic acid ethyl ester (retigabine). The effect of retigabine persisted in the presence of tetrodotoxin and simultaneous blockade of GABAA receptors, small-conductance calcium-activated K+ (SK) channels, and hyperpolarization-activated (Ih) channels, and it was potently reversed by the KCNQ channel blocker 4-pyridinylmethyl-9(10H)-anthracenone (XE991), indicating a direct effect on KCNQ channels. Likewise, in vivo single unit recordings from dopaminergic neurons revealed a prominent reduction in spike activity after systemic administration of retigabine. Furthermore, retigabine inhibited dopamine synthesis and c-Fos expression in the striatum under basal conditions. Retigabine completely blocked the excitatory effect of dopamine D2 autoreceptor antagonists. Again, the in vitro and in vivo effects of retigabine were completely reversed by preadministration of XE991. Dual immunocytochemistry revealed that KCNQ4 is the major KCNQ channel subunit expressed in all dopaminergic neurons in the mesolimbic and nigrostriatal pathways. Collectively, these observations indicate that retigabine negatively modulates dopaminergic neurotransmission, likely originating from stimulation of mesencephalic KCNQ4 channels.

Hydrogen peroxide hyperpolarizes rat CA1 pyramidal neurons by inducing an increase in potassium conductance.

It has been suggested that hydrogen peroxide is involved in cascades of pathological events affecting neural cells. The aim of this study was therefore to examine whether this molecule is able by itself to modify membrane properties of pyramidal neurons in the CA1 region of the rat hippocampus. Intracellular recordings in the slice preparation showed that 3.3 mM hydrogen peroxide hyperpolarized all neurons tested (n = 41) by 11 +/- 3 mV. This effect persisted in the presence of tetrodotoxin. It developed slowly, was reversible and reproducible. In the presence of tetrodotoxin, the extrapolated reversal potential of this effect was -95 +/- 5 mV in 2.5 mM external potassium. This value was not significantly different from the one obtained with the GABAB agonist baclofen (10 microM) (-98 +/- 5 mV). It shifted when the concentration of external potassium was increased to 10.5 mM (from -96 +/- 5 to -62 +/- 4 mV), in close agreement with the Nernst equation potassium ions. The hyperpolarization was significantly reduced (by 65 +/- 22%) by the potassium channel blocker barium (100 microM). We suggest that hydrogen peroxide is able to induce an increase in potassium conductance in rat CA1 pyramidal neurons. The exact mechanism by which it produces this effect (direct action on channels or indirect effect) remains to be determined.

Inhibition of in vitro and ex vivo uptake of noradrenaline and 5-hydroxytryptamine by five antidepressants; Correlation with reduction of spontaneous firing rate of central monoaminergic neurones

SummaryThe principal neurochemical property of tricyclic antidepressants is the blockade of noradrenaline (NA) and/or 5-hydroxytryptamine (5-HT) uptake into monoaminergic nerve endings. Electrophysiological studies show that these drugs also decrease the firing rate of the noradrenergic neurones of the locus coeruleus (L.C.) and of the serotonergic neurones of the dorsal raphe (D.R.). In order to assess the relation between the two phenomena, the influence of five tricyclic antidepressants on NA and 5-HT uptake was studied in vitro. The concentrations required to produce a 50% inhibition (IC50) were determined and correlated with the respective doses required to reduce to 50% (ID50) the firing rate of L.C. and D.R. neurones. Ex vivo experiments were also performed to study the influence of the tricyclic antidepressants on NA and 5-HT uptake when administered i.v. at the doses decreasing to 50% the firing rate of L.C. and D.R. cells.The inhibition of the NA uptake by tricyclic antidepressants can account, at least in part, for the inhibition of the firing rate of L.C. neurones observed after acute i.v. administration. In the case of serotonergic neurons, the results do not allow a firm conclusion.

SK Channel blockade promotes burst firing in dorsal raphe serotonergic neurons

Previous in vivo studies have shown that blockade of small‐conductance Ca2+‐activated potassium (SK) channels enhances burst firing in dopaminergic neurons. As bursting has been found to be physiologically relevant for the synaptic release of serotonin (5‐HT), we investigated the possible role of SK channels in the control of this firing pattern in 5‐HT neurons of the dorsal raphe nucleus. In these cells, bursts are usually composed of doublets consisting of action potentials separated by a small interval (< 20 ms). Both in vivo and in vitro extracellular recordings were performed, using anesthetized rats and rat brain slices, respectively. In vivo, the specific SK blocker UCL 1684 (200 μm) iontophoresed onto presumed 5‐HT neurons significantly increased the production of bursts in 13 out of 25 cells. Furthermore, the effect of UCL 1684 persisted in the presence of both the GABAA antagonist SR 95531 (10 mm) and the GABAB antagonist CGP 35348 (10 mm), whereas these agents by themselves did not significantly influence the neuronal firing pattern. In vitro, bath superfusion of the SK channel blocker apamin (300 nm) induced bursting in only three out of 18 neurons, although it increased the coefficient of variation of the interspike intervals in all the other cells. Our results suggest that SK channel blockade promotes bursting activity in 5‐HT neurons via a direct action. An input which is present only in vivo seems to be important for the induction of this firing pattern in these cells.

Electrophysiological characterization of the SK channel blockers methyl-laudanosine and methyl-noscapine in cell lines and rat brain slices.

We have recently shown that the alkaloid methyl‐laudanosine blocks SK channel‐mediated afterhyperpolarizations (AHPs) in midbrain dopaminergic neurones. However, the relative potency of the compound on the SK channel subtypes and its ability to block AHPs of other neurones were unknown. Using whole‐cell patch‐clamp experiments in transfected cell lines, we found that the compound blocks SK1, SK2 and SK3 currents with equal potency: its mean IC50s were 1.2, 0.8 and 1.8 μM, respectively. IK currents were unaffected. In rat brain slices, methyl‐laudanosine blocked apamin‐sensitive AHPs in serotonergic neurones of the dorsal raphe and noradrenergic neurones of the locus coeruleus with IC50s of 21 and 19 μM, as compared to 15 μM in dopaminergic neurones. However, at 100 μM, methyl‐laudanosine elicited a constant hyperpolarization of serotonergic neurones of about 9 mV, which was inconsistently (i.e. not in a reproducible manner) antagonized by atropine and hence partly due to the activation of muscarinic receptors. While exploring the pharmacology of related compounds, we found that methyl‐noscapine also blocked SK channels. In cell lines, methyl‐noscapine blocked SK1, SK2 and SK3 currents with mean IC50s of 5.9, 5.6 and 3.9 μM, respectively. It also did not block IK currents. Methyl‐noscapine was slightly less potent than methyl‐laudanosine in blocking AHPs in brain slices, its IC50s being 42, 37 and 29 μM in dopaminergic, serotonergic and noradrenergic neurones, respectively. Interestingly, no significant non‐SK effects were observed with methyl‐noscapine in slices. At a concentration of 300 μM, methyl‐noscapine elicited the same changes in excitability in the three neuronal types than did a supramaximal concentration of apamin (300 nM). Methyl‐laudanosine and methyl‐noscapine produced a rapidly reversible blockade of SK channels as compared with apamin. The difference between the IC50s of apamin (0.45 nM) and methyl‐laudanosine (1.8 μM) in SK3 cells was essentially due to a major difference in their k−1 (0.028 s−1 for apamin and 20 s−1 for methyl‐laudanosine). These experiments demonstrate that both methyl‐laudanosine and methyl‐noscapine are medium potency, quickly dissociating, SK channel blockers with a similar potency on the three SK subtypes. Methyl‐noscapine may be superior in terms of specificity for the SK channels.

M-type channels selectively control bursting in rat dopaminergic neurons

Midbrain dopaminergic neurons in the substantia nigra, pars compacta and ventral tegmental area are critically important in many physiological functions. These neurons exhibit firing patterns that include tonic slow pacemaking, irregular firing and bursting, and the amount of dopamine that is present in the synaptic cleft is much increased during bursting. The mechanisms responsible for the switch between these spiking patterns remain unclear. Using both in‐vivo recordings combined with microiontophoretic or intraperitoneal drug applications and in‐vitro experiments, we have found that M‐type channels, which are present in midbrain dopaminergic cells, modulate the firing during bursting without affecting the background low‐frequency pacemaker firing. Thus, a selective blocker of these channels, 10,10‐bis(4‐pyridinylmethyl)‐9(10H)‐anthracenone dihydrochloride, specifically potentiated burst firing. Computer modeling of the dopamine neuron confirmed the possibility of a differential influence of M‐type channels on excitability during various firing patterns. Therefore, these channels may provide a novel target for the treatment of dopamine‐related diseases, including Parkinson’s disease and drug addiction. Moreover, our results demonstrate that the influence of M‐type channels on the excitability of these slow pacemaker neurons is conditional upon their firing pattern.

New assessment of dependency in demented patients: impact on the quality of life in informal caregivers.

Abstract  A qualitative tool was recently developed for evaluation of dependency in a demented population. This tool assesses the impact of cognitive impairment on functional status, taking into account disability in both the basic and the instrumental activities of daily living. The purpose of the present paper was to study the impact of dependency on informal caregivers who assist demented patients at home, with this new useful tool. Methods: A cross‐sectional analysis was undertaken of the subgroup of 145 demented patients of the National Dementia Economic Study, aged ≥65 years, living in the community, with an available caregiver. A neuropsychological assessment of patients (Mini‐Mental State Examination) and a comprehensive evaluation of caregivers (quality of life, Short Form Health Survey‐36, depression, Sense of Competence) were recorded. A total of 32.4% were dependent, disabled in both basic and instrumental functions, 42.1% were non‐dependent but with instrumental functional disabilities and 25.5% were non‐dependent. Impact of dependency on the caregivers experience was significant for different aspects (satisfaction with caregiving, subjective burden, quality of life, depression). Medical and non‐medical costs increased with the severity of functional disability. Findings indicate that this tool is also useful to assess the impact of progression of functional disability in patients with dementia, on the caregiver issues. The consequences appeared both on personal feelings and on quality of life and financial involvement in management of the patient. Cognitive impairment appears to have more moderate repercussions in these areas.