Nitroxyl (HNO) is a redox sibling of nitric oxide (NO) that targets distinct signalling pathways with pharmacological endpoints of high significance in the treatment of heart failure. Beneficial HNO effects depend, in part, on its ability to release calcitonin gene-related peptide (CGRP) through an unidentified mechanism. Here we propose that HNO is generated as a result of the reaction of the two gasotransmitters NO and H2S. We show that H2S and NO production colocalizes with transient receptor potential channel A1 (TRPA1), and that HNO activates the sensory chemoreceptor channel TRPA1 via formation of amino-terminal disulphide bonds, which results in sustained calcium influx. As a consequence, CGRP is released, which induces local and systemic vasodilation. H2S-evoked vasodilatatory effects largely depend on NO production and activation of HNO–TRPA1–CGRP pathway. We propose that this neuroendocrine HNO–TRPA1–CGRP signalling pathway constitutes an essential element for the control of vascular tone throughout the cardiovascular system.

Calcitonin gene-related peptide (CGRP) is a key mediator in primary headaches including migraine. Animal models of meningeal nociception demonstrate both peripheral and central CGRP effects; however, the target structures remain unclear. To study the distribution of CGRP receptors in the rat trigeminovascular system we used antibodies recognizing two components of the CGRP receptor, the calcitonin receptor-like receptor (CLR) and the receptor activity-modifying protein 1 (RAMP1). In the cranial dura mater, CLR and RAMP1 immunoreactivity (-ir) was found within arterial blood vessels, mononuclear cells, and Schwann cells, but not sensory axons. In the trigeminal ganglion, besides Schwann and satellite cells, CLR- and RAMP1-ir was found in subpopulations of CGRP-ir neurons where colocalization of CGRP- and RAMP1-ir was very rare ( approximately 0.6%). CLR- and RAMP1-ir was present on central, but not peripheral, axons. In the spinal trigeminal nucleus, CLR- and RAMP1-ir was localized to "glomerular structures," partly colocalized with CGRP-ir. However, CLR- and RAMP1-ir was lacking in central glia and neuronal cell bodies. We conclude that CGRP receptors are associated with structural targets of known CGRP effects (vasodilation, mast cell degranulation) and targets of unknown function (Schwann cells). In the spinal trigeminal nucleus, CGRP receptors are probably located on neuronal processes, including primary afferent endings, suggesting involvement in presynaptic regulation of nociceptive transmission. Thus, in the trigeminovascular system CGRP receptor localization suggests multiple targets for CGRP in the pathogenesis of primary headaches.

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Migraine is a complex neurological disease with a genetic background. Headache is the most prominent and clinically important symptom of migraine but its origin is still enigmatic. Numerous clinical, histochemical, electrophysiological, molecular and genetical approaches form a puzzle of findings that slowly takes shape. The generation of primary headaches like migraine pain seems to be the consequence of multiple pathophysiological changes in meningeal tissues, the trigeminal ganglion, trigeminal brainstem nuclei and descending inhibitory systems, based on specific characteristics of the trigeminovascular system. This contribution reviews the current discussion of where and how the migraine pain may originate and outlines the experimental work to answer these questions.

Axonal conduction velocity varies according to the level of preceding impulse activity. In unmyelinated axons this typically results in a slowing of conduction velocity and a parallel increase in threshold. It is currently held that Na + -K + -ATPase-dependent axonal hyperpolarization is responsible for this slowing but this has long been equivocal. We therefore examined conduction velocity changes during repetitive activation of single unmyelinated axons innervating the rat cranial meninges. In direct contradiction to the currently accepted postulate, Na + -K + -ATPase blockade actually enhanced activity-induced conduction velocity slowing, while the degree of velocity slowing was curtailed in the presence of lidocaine (10-300 μM) and carbamazepine (30-500 μM) but not tetrodotoxin (TTX, 10-80 nM). This suggests that a change in the number of available sodium channels is the most prominent factor responsible for activity-induced changes in conduction velocity in unmyelinated axons. At moderate stimulus frequencies, axonal conduction velocity is determined by an interaction between residual sodium channel inactivation following each impulse and the retrieval of channels from inactivation by a concomitant Na + -K + -ATPase-mediated hyperpolarization. Since the process is primarily dependent upon sodium channel availability, tracking conduction velocity provides a means of accessing relative changes in the excitability of nociceptive neurons.

Ultrastructural, immunocytochemical, and immunoelectron microscopical examinations are reported that describe the morphology of putative sensory nerve endings in the dura mater encephali of the rat and the cat. Morphometrical measurements and reconstructions showed that in the cat the mean diameter of axons, the bare area of axolemma, and the content of mitochondria and vesicles are highly variable in dural nerve endings. Nerve fibers with a high volume density of mitochondria are thought to be sensory, while nerve fibers containing many small vesicles are considered autonomic. There is, however, a broad overlap of mitochondria-rich and vesicle-rich nerve fibers in the dura, so that discrimination between sensory and autonomic endings by these characteristics frequently fails. Whole-mount preparations treated cytochemically for detection of substance P- and calcitonin gene-related peptide-like immunoreactivity in the rat and the cat showed a network of immunopositive nerve fibers in the vicinity of dural blood vessels. Most of these peptidergic and probably sensory nerve fibers were found terminating in the dural connective tissue far from vessels. Calcitonin gene-related peptide-positive nerve fibers were much more abundant than substance P-positive fibers. Immunoelectron microscopic preparations revealed that calcitonin gene-related peptide- and substance P-like immunoreactivity is found in a small proportion of generally thin unmyelinated nerve fibers. These proportions were very similar in the rat and the cat. Summarizing the recent literature, the morphological characteristics of putative sensory nerve fibers in the dura mater are discussed in relation to their possible functional significance for neurogenic inflammation and nociception.

Japan1 The dura mater encephali of the rat was exposed and the blood flow around branches of the medial meningeal artery was monitored with a laser Doppler flowmeter. Changes in the meningeal blood flow (MBF) following electrical stimulation of the dura mater at a parasagittal site were registered. The effects of human calcitonin gene-related peptide (h-aCGRP) and the CGRP antagonist (h-aCGRP8-37) on the MBF were tested. 2 Electrical stimulation with rectangular pulses of 0.5 ms, 10-20 V, 5-10 Hz and a duration of 30 s caused an increase of the MBF in 14 out of 16 rats tested. The increases were dependent on stimulus strength and frequency. 3 The increase in MBF was inhibited in a dose-dependent manner by topical application of 0.1 ml of h-mCGRP837 at concentrations of 10-7-10-'M. The highest dose abolished the increase in MBF. 4 Topical administration of 0.1 ml of h-aCGRP at a concentration of 10-4 M increased the basal MBF by 15% on average. 5 It is suggested that the increase in MBF following electrical stimulation of the dura mater is mediated by the release of CGRP. The contribution of the dural afferent and sympathetic and parasympathetic efferent nerve fibres to this response are discussed.

Heat sensitivity shows considerable functional variability in humans and laboratory animals, and is fundamental to inflammatory and possibly neuropathic pain. In the mouse, at least, much of this variability is genetic because inbred strains differ robustly in their behavioral sensitivity to noxious heat. These strain differences are shown here to reflect differential responsiveness of primary afferent thermal nociceptors to heat stimuli. We further present convergent behavioral and electrophysiological evidence that the variable responses to noxious heat are due to strain-dependence of CGRP expression and sensitivity. Strain differences in behavioral response to noxious heat could be abolished by peripheral injection of CGRP, blockade of cutaneous and spinal CGRP receptors, or long-term inactivation of CGRP with a CGRP-binding Spiegelmer. Linkage mapping supports the contention that the genetic variant determining variable heat pain sensitivity across mouse strains affects the expression of the Calca gene that codes for CGRP␣.calcitonin gene-related peptide ͉ genetic ͉ Calca ͉ nociceptors ͉ pain H umans display wide individual variability in sensitivity to pain.Although the relative importance of genes versus experience in human pain perception is unclear, recent studies have shown that mouse strains display large differences in behavioral pain sensitivity that are heritable (1). These same studies revealed genetic correlations between baseline thermal nociception and the hypersensitivity states associated with inflammatory and neuropathic pain (2). Of the strains examined, AKR and C57BL͞6 mice displayed the largest and most consistent differences in several different assays of thermal nociception, with AKR being much less sensitive than C57BL͞6. In contrast, AKR mice exhibit more robust heat hyperalgesia after inflammatory or nerve injury (1). Despite our considerable knowledge of the behavioral ''phenomics'' of baseline heat pain and hyperalgesia, there are almost no published data regarding the underlying cellular or molecular mechanisms. Here we show that the observed strain differences in response to thermal stimulation are caused by corresponding differences in the functioning of primary afferent nociceptors. The differences in nociceptor sensitivity, in turn, are caused by the presence of and sensitivity to the neuropeptide calcitonin gene-related polypeptide (CGRP). Finally, linkage mapping revealed a candidate gene likely responsible for the strain difference: Calca, the gene encoding CGRP␣.CGRP␣ is a secretory neuropeptide released from thin nerve fibers at their peripheral and central terminals, which is thought to contribute importantly to neurogenic inflammation in the skin and to central sensitization in the spinal cord (3-6). CGRP acts through a G s protein-coupled receptor complex to activate cAMPdependent protein kinase (PKA). PKA, via the transcription factor cAMP response element-binding protein, enhances expression of pronociceptive genes including the Calca gene itself (7-9). Moreover, many...