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

Abstract

1 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. 2 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 IC 50 s were 1.2, 0.8 and 1.8 mM, respectively. IK currents were unaffected. In rat brain slices, methyl-laudanosine blocked apaminsensitive AHPs in serotonergic neurones of the dorsal raphe and noradrenergic neurones of the locus coeruleus with IC 50 s of 21 and 19 mM, as compared to 15 mM in dopaminergic neurones. However, at 100 mM, 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. 3 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 IC 50 s of 5.9, 5.6 and 3.9 mM, respectively. It also did not block IK currents. Methyl-noscapine was slightly less potent than methyl-laudanosine in blocking AHPs in brain slices, its IC 50 s being 42, 37 and 29 mM 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 mM, methyl-noscapine elicited the same changes in excitability in the three neuronal types than did a supramaximal concentration of apamin (300 nM). 4 Methyl-laudanosine and methyl-noscapine produced a rapidly reversible blockade of SK channels as compared with apamin. The difference between the IC 50 s of apamin (0.45 nM) and methyllaudanosine (1.8 mM) in SK3 cells was essentially due to a major difference in their k À1 (0.028 s À1 for apamin and X20 s À1 for methyl-laudanosine). 5 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.

Functions and Modulation of Neuronal SK Channels

“…More recently, the reversible blockers N-methyl-laudanosine (NML) and methyl-noscapine [40,41], bis-quaternary isoquinoliniums [42] and lipophilic tetrahydroisoquinoline derivatives [43] have been developed. Tamapin from scorpion venom [44] and Lei-Dab7, a synthetic derivative of Leiurotoxin [45] selectively block K Ca 2.2 channels.…”

“…SK channels control the firing patterns of thalamocortical relay neurons [73], subthalamic neurons [68], globus pallidus neurons [75], cerebellar Purkinje neurons [69,70], deep cerebellar nuclei neurons (which express K Ca 2.1 and K Ca 2.2 channels; [71]), dorsal vagal neurons (mediated by K Ca 2.3 channels; [20,65]), mitral cells in the olfactory bulb [72], neurons in the inferior olive [91,92], neurons in the preBötzinger complex [74], central vestibular nuclei neurons [93], sympathetic neurons [66], noradrenergic neurons in the locus coreuleus [40], and spinal and hypoglossal motoneurons [94,95].…”

“…The medium AHP (mAHP) activates rapidly and decays over several hundred milliseconds, and is blocked by apamin in layer 5 pyramidal cortical neurons [63], vagal neurons [64,65], sympathetic neurons [66], dopaminergic neurons in the substantia nigra [22], lateral [62] and basolateral [67] amygdala pyramidal neurons, subthalamic neurons [68], cerebellar Purkinje neurons [69,70], deep cerebellar nuclei neurons [71], serotonergic neurons in the dorsal raphe [40], noradrenergic neurons in the locus coeruleus [40], mitral cells in the olfactory bulb [72], thalamocortical relay neurons [73], neurons in the pre-Bötzinger complex that are involved in respiratory rhythm generation [74] and globus pallidus neurons [75], showing that SK channels contribute to the mAHP in all of these brain regions.…”

Physiologie, pharmacologie et modélisation de canaux potassiques

“…Ce programme a abouti à la découverte d'autres composés quaternaires, la N-méthyl-laudanosine (NML) et la N-méthyl-noscapine (NMN) ( Figure 5). Ces molécules bloquent les courants SK avec une puissance moyenne (IC 50 ≈ 2-4 μM) et de manière non sélective [41,42]. La faible puissance de la NML est due à un k off plus élevé que celui de l'apamine, ce qui en fait un bloqueur plus facilement réversible [41].…”

“…Ces molécules bloquent les courants SK avec une puissance moyenne (IC 50 ≈ 2-4 μM) et de manière non sélective [41,42]. La faible puissance de la NML est due à un k off plus élevé que celui de l'apamine, ce qui en fait un bloqueur plus facilement réversible [41]. Le développement de dimères dérivés de la NML a ensuite permis de synthétiser des bloqueurs SK plus puissants dont l'affinité est proche de celle de l'UCL 1684 (Ki ≈ 20 nM) [43].…”