Chemical Determinants Involved in Anandamide-induced Inhibition of T-type Calcium Channels

Abstract

Anandamide, originally described as an endocannabinoid, is the main representative molecule of a new class of signaling lipids including endocannabinoids and N-acyl-related molecules, eicosanoids, and fatty acids. Bioactive lipids regulate neuronal excitability by acting on G-protein-coupled receptors (such as CB1) but also directly modulate various ionic conductances including voltage-activated T-type calcium channels (T-channels). However, little is known about the properties and the specificity of this new class of molecules on their various targets. In this study, we have investigated the chemical determinants involved in anandamide-induced inhibition of the three cloned T-channels: Ca V 3.1, Ca V 3.2, and Ca V 3.3. We show that both the hydroxyl group and the alkyl chain of anandamide are key determinants of its effects on T-currents. As follows, T-currents are also inhibited by fatty acids. Inhibition of the three Ca V 3 currents by anandamide and arachidonic acid does not involve enzymatic metabolism and occurs in cell-free inside-out patches. Inhibition of T-currents by fatty acids and N-acyl ethanolamides depends on the degree of unsaturation but not on the alkyl chain length and consequently is not restricted to eicosanoids. Inhibition increases for polyunsaturated fatty acids comprising 18 -22 carbons when cis-double bonds are close to the carboxyl group. Therefore the major natural (food-supplied) and mammalian endogenous fatty acids including ␥-linolenic acid, mead acid, and arachidonic acid as well as the fully polyunsaturated 3-fatty acids that are enriched in fish oil eicosapentaenoic and docosahexaenoic acids are potent inhibitors of T-currents, which possibly contribute to their physiological functions.Voltage-dependent calcium channels comprise three families: the L-type channels (Ca V 1 family), the neuronal N-, P/Q-, and R-type channels (Ca V 2 family), and the T-type channels (Ca V 3 family). The electrophysiological features of T-type Ca 2ϩ channels (T-channels) are low voltage-activated Ca 2ϩ currents, low unitary conductance, fast inactivation and slow deactivation kinetics, and strong steady-state inactivation at physiological resting potentials (1, 2). Three T-channel subunits have been identified: Ca V 3.1 (or ␣ 1G ), Ca V 3.2 (or ␣ 1H ), and Ca V 3.3 (or ␣ 1I ) (2). In mammalian expression systems, the Ca V 3.1 and Ca V 3.2 currents share typical properties of native T-channels, while Ca V 3.3 currents display unusually slow inactivation kinetics, which are characteristic of specific neurons (1-3). These three subunits are also subject to alternative splicing, which influences their electrophysiological properties (4 -6). The Ca V 3.1 and Ca V 3.2 subunits are widely expressed in various tissues, mainly in the heart and in the nervous system, whereas Ca V 3.3 is restricted to the central nervous system (2). In the nervous system, T-channels generate low threshold spikes and participate in spontaneous firing (1, 2). They are involved in slow wave sleep (7), in absence epilepsy (8, 9),...

N-Arachidonyl Glycine Does Not Activate G Protein–Coupled Receptor 18 Signaling via Canonical Pathways

“…3 and 4). Similar effects of lipoamino acids and arachidonic acid, a product of fatty acid amide hydrolase (FAAH) hydrolysis of NAGly (Grazia Cascio et al, 2004), on ion channel function have been documented (Chemin et al, 2007;Guo et al, 2008a;Barbara et al, 2009). However, it is unlikely that the effects observed in this study are a result of NAGly breakdown to arachidonic acid and glycine because arachidonic acid reduces N-type Ca 21 channel amplitude (Liu and Rittenhouse, 2000) and the effect of NAGly on T-type Ca 21 channels is independent of FAAH activity (Barbara et al, 2009).…”

International Union of Basic and Clinical Pharmacology. LXXIX. Cannabinoid Receptors and Their Ligands: Beyond CB1and CB2

“…Certain cannabinoid receptor agonists (Table 7) and antagonists have been found to antagonize T-type voltage-gated calcium channels at concentrations in the mid-nanomolar or low micromolar range. Thus, for example, evidence has been obtained (Chemin et al, 2001(Chemin et al, , 2007Ross et al, 2008Ross et al, , 2009 It is noteworthy that the phytocannabinoid cannabidiol has also been found to inhibit cloned Ca V 3.1, Ca V 3.2, and Ca V 3.3 calcium channels, its reported IC 50 values for this inhibition being 813, 776, and 3631 nM, respectively (Ross et al, 2008). In addition, a recent article reported that the endogenous lipo-amino acids, NAGly and N-arachidonoyl GABA, can potently inhibit Ca V 3.1, Ca V 3.2, and Ca V 3.3 channels (IC 50 Ͻ1.0 M) (Barbara et al, 2009).…”

Cross-Modulation and Molecular Interaction at the Cav3.3 Protein between the Endogenous Lipids and the T-Type Calcium Channel Antagonist TTA-A2.

“…These molecules include arachidonic acid, v3-fatty acids, endocannabinoids (as anandamide), lipo-amino-acids and lipo-neurotransmitters (Zhang et al, 2000;Chemin et al, 2001Chemin et al, , 2007Talavera et al, 2004;Danthi et al, 2005;Barbara et al, 2009;Ross et al, 2009;Gilmore et al, 2012). These lipids are implicated in multiple physiologic functions and more specifically in pain perception (Bradshaw and Walker, 2005;Burstein, 2008;Basbaum et al, 2009), sleep and epilepsy (Chen and Bazan, 2005), and heart rhythm and vasodilatation (Roman, 2002;Leaf et al, 2003).…”