arachidonyl-dopamine and Pain

Mass spectrometric approaches to the identification and quantification of lipid signalling molecules are reviewed. Fatty acid amides are an important new class of lipid signalling molecules which include oleamide, the endocannabinoid anandamide, the endovanilloid/endocannabinoid N-arachidonoyldopamine (NADA) and the endovanilloid N-oleoyldopamine (OLDA) among many others. This diverse group of endogenous compounds comprises combinations of acyl backbones coupled by an amide bond to any of a variety of different small polar molecules such as ethanolamine, various amino acids, and catecholamines. Many fatty acid amides appear to play a role in pain and inflammation. Targeted lipidomics of fatty acid amides aims to identify new members of this diverse class of compounds, of which only a few representative molecules have been characterized to date. This effort has been made feasible by advances in chromatography and mass spectrometry, which permits: (1) identification of compounds present in complex mixtures, (2) astronomical increases in sensitivity due to miniaturization of HPLC components, and (3) novel scanning modes that permit the identification of compounds exhibiting similar structural components. Insofar as lipid signalling molecules such as prostanoids, leukotrienes and endocannabinoids operate via G-protein coupled receptors (GPCR), it appears likely that many of the numerous lipids awaiting identification may serve as ligands for any of the greater than 150 orphan GPCRs.

Topics: Amides; Analgesics; Animals; Arachidonic Acids; Chromatography, High Pressure Liquid; Dopamine; Fatty Acids; Humans; Inflammation; Lipid Metabolism; Lipids; Mass Spectrometry; Models, Chemical; Pain; Receptors, G-Protein-Coupled; Signal Transduction

Transient receptor potential vanilloid receptor 1 (TRPV1)-mediated release of neuroactive peptides and neurotransmitters from the peripheral and central terminals of primary sensory neurons can critically contribute to nociceptive processing at the periphery and in the CNS. However, the mechanisms that link TRPV1 activation with Ca2+ signaling at the release sites and neurosecretion are poorly understood. Here we demonstrate that a brief stimulation of the receptor using either capsaicin or the endogenous TRPV1 agonist N-arachidonoyl-dopamine induces a prolonged elevation of presynaptic [Ca2+](i) and a concomitant enhancement of glutamate release at sensory synapses. Initiation of this response required Ca2+ entry, primarily via TRPV1. The sustained phase of the response was independent of extracellular Ca2+ and was prevented by inhibitors of mitochondrial Ca2+ uptake and release mechanisms. Measurements using a mitochondria-targeted Ca2+ indicator, mtPericam, revealed that TRPV1 activation elicits a long-lasting Ca2+ elevation in presynaptic mitochondria. The concentration of TRPV1 agonist determined the duration of mitochondrial and cytosolic Ca2+ signals in presynaptic boutons and, consequently, the period of enhanced glutamate release and action potential firing by postsynaptic neurons. These data suggest that mitochondria control vanilloid-induced neurotransmission by translating the strength of presynaptic TRPV1 stimulation into duration of the postsynaptic response.

Topics: Action Potentials; Afferent Pathways; Animals; Animals, Newborn; Arachidonic Acids; Calcium; Calcium Signaling; Capsaicin; Cells, Cultured; Coculture Techniques; Dopamine; Ganglia, Spinal; Glutamic Acid; Mitochondria; Neurons, Afferent; Nociceptors; Pain; Patch-Clamp Techniques; Posterior Horn Cells; Presynaptic Terminals; Rats; Rats, Sprague-Dawley; Spinal Nerve Roots; Synaptic Transmission; TRPV Cation Channels

Multiple lines of evidence suggest that calcium/calmodulin-dependent kinase II alpha (CaMKIIalpha) plays an important role in the spinal dorsal horn in nociceptive models of chemical, inflammatory and nerve injury. Moreover, CaMKIIalpha phosphorylates the vanilloid receptor type 1 (TRPV1), thereby regulating vanilloid agonist binding to the receptor. Herein, we have explored a possible interaction of CaMKIIalpha activity with the TRPV1 receptor in rat trigeminal ganglion (TG) neurons in vitro. Inhibition of CaMKIIalpha with KN-93 (5 microM) inhibited capsaicin (CAP)- and n-arachidonoyl-dopamine (NADA)-evoked calcitonin gene-related peptide (CGRP) release effectively decreasing the Emax for both compounds. This effect was not mimicked by the inactive compound KN-92 (5 microM), indicating that the effect was mediated by CaMKIIalpha inhibition. CAP also stimulated a significant approximately 50% increase in autophosphorylation of CaMKIIalpha at Thr286/287. Immunocytochemistry for phospho-CaMKIIalpha indicated that this effect specifically occurred in TRPV1-positive TG neurons. These findings indicate that phopho-CaMKIIalpha is likely to play a role in presynaptic primary afferents in animal models of nociceptive hypersensitivity and provide support for CaMKIIalpha modulation of TRPV1 activity in sensory neurons.

Topics: Animals; Arachidonic Acids; Calcitonin Gene-Related Peptide; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases; Capsaicin; Cells, Cultured; Dopamine; Dose-Response Relationship, Drug; Enzyme Inhibitors; Neurons, Afferent; Nociceptors; Pain; Phosphorylation; Rats; Rats, Sprague-Dawley; Threonine; Trigeminal Ganglion; TRPV Cation Channels