Marlene L. Cohen

5-HT1F receptor agonists inhibit neurogenic dural inflammation in guinea pigs.

THE serotonin (5-HT) receptor subtype mediating inhibition of neurogenic dural inflammation in guinea pigs was investigated using a series of serotonin agonists with differing affinities for the 5-HT1B, 5-HT1D and 5-HT1F receptors. When agonist potencies for inhibiting neurogenic inflammation were compared with affinities for these receptor subtypes, a significant positive correlation was seen only with the 5-HT1F receptor. The potency of agonists in inhibiting adenylate cyclase in cells transfected with human 5-HT1F receptor was also highly correlated with their potency in the animal model of migraine. In situ hybridization demonstrated 5-HT1F receptor mRNA in guinea pig trigeminal ganglion neurons. These data suggest that the 5-HT1F receptor is a rational target for migraine therapeutics.

Serotonin in migraine : theories, animal models and emerging therapies

A role for serotonin in migraine has been supported by changes in circulating levels of serotonin and its metabolites during the phases of a migraine attack, along with the ability of serotonin-releasing agents to induce migraine-like symptoms. The development of serotonin receptor agonists with efficacy in the clinic for the alleviation of migraine pain further implicates serotonin as a key molecule in migraine. Several theories regarding the etiology of migraine have been proposed. The vasodilatory theory of migraine suggested that extracranial arterial dilation during an attack was related to migraine pain; a theory supported when vasoconstrictors such as sumatriptan alleviated migraine pain. The neurological theory of migraine proposed that migraine resulted from abnormal firing in brain neurons. Cortical spreading depression, one facet of the neurological theory, could explain the prodrome of migraine. The neurogenic dural inflammation theory of migraine supposed that the dural membrane surrounding the brain became inflamed and hypersensitive due to release of neuropeptides from primary sensory nerve terminals. Substance P, calcitonin gene related peptide and nitric oxide are all though to play a role in the dural inflammatory cascade. Animal models of migraine have been utilized to study the physiology of migraine and develop new pharmaceutical therapies. One model measures the shunting of blood to arteriovenous anastomoses based on a proposal that migraine primarily involves cranial arteriovenous vasodilation. Another model utilizes electrical stimulation of the trigeminal ganglion to induce neurogenic dural inflammation quantified by the resulting extravasation of proteins. Pharmacological agents such as meta-chlorophenylpiperazine (mCPP) and nitroglycerin have also been used to induce dural extravasation in animals. Both compounds also induce migraine attacks in individuals with a history of migraine. In addition, Fos, a protein produced by activation of the c-fos gene, has been measured as an index of migraine-like pain transmission to the CNS following chemical or electrical stimulation of the trigeminal nerve. A role for serotonin in migraine is further supported by the efficacy of serotonin receptor ligands. Sumatriptan is an agonist at 5-HT1D and 5-HT1B receptor subtypes, and effective in treating migraine pain and associated symptoms. Recently, selective 5-HT1F agonists have been proposed for the treatment of migraine, without the side effects associated with the present 5-HT1D and 5-HT1B receptor agonists. A role for 5-HT2B receptors has also been suggested the initiation of migraine, supporting use of selective 5-HT2B receptor antagonists in migraine. Thus, agents that modulate 5-HT1B, 5-HT1D, 5-HT1F and 5-HT2B receptors either have or may have clinical utility in the therapy of migraine headache.

Somatostatin inhibits adrenergic and cholinergic neurotransmission in smooth muscle.

Abstract Somatostatin reduced the response to field stimulation in the guinea pig ileum and reduced the spontaneous contractions in the rabbit jejunum, an effect that was blocked by tetrodotoxin. Somatostatin also inhibited field stimulated alpha adrenergic contractions in the rat vas deferens and rabbit ear artery. However, the responses to direct application of either acetylcholine in the ileum or to norepinephrine in the ear artery or vas deferens were not affected by somatostatin. These results strongly suggest that somatostatin inhibits neuronal release of cholinergic and adrenergic transmitter substances in smooth muscle.

Preclinical pharmacological profile of the selective 5-HT1F receptor agonist lasmiditan

Introduction: Lasmiditan (also known as COL-144 and LY573144; 2,4,6-trifluoro-N-[6-[(1-methylpiperidin-4-yl)carbonyl]pyridin-2yl]benzamide) is a high-affinity, highly selective serotonin (5-HT) 5-HT1F receptor agonist. Results: In vitro binding studies show a Ki value of 2.21 nM at the 5-HT1F receptor, compared with Ki values of 1043 nM and 1357 nM at the 5-HT1B and 5-HT1D receptors, respectively, a selectivity ratio greater than 470-fold. Lasmiditan showed higher selectivity for the 5-HT1F receptor relative to other 5-HT1 receptor subtypes than the first generation 5-HT1F receptor agonist LY334370. Unlike the 5-HT1B/1D receptor agonist sumatriptan, lasmiditan did not contract rabbit saphenous vein rings, a surrogate assay for human coronary artery constriction, at concentrations up to 100 µM. In two rodent models of migraine, oral administration of lasmiditan potently inhibited markers associated with electrical stimulation of the trigeminal ganglion (dural plasma protein extravasation, and induction of the immediate early gene c-Fos in the trigeminal nucleus caudalis). Conclusions: Lasmiditan presents a unique pyridinoyl-piperidine scaffold not found in any other antimigraine class. Its chemical structure and pharmacological profile clearly distinguish it from the triptans. The potency and selectivity of lasmiditan make it ideally suited to definitively test the involvement of 5-HT1F receptors in migraine headache therapy.

Pharmacological Characterization of the Competitive GLUK5 Receptor Antagonist Decahydroisoquinoline LY466195 in Vitro and in Vivo

The excitatory neurotransmitter glutamate has been implicated in both migraine and persistent pain. The identification of the kainate receptor GLUK5 in dorsal root ganglia, the dorsal horn, and trigeminal ganglia makes it a target of interest for these indications. We examined the in vitro and in vivo pharmacology of the competitive GLUK5-selective kainate receptor antagonist LY466195 [(3S,4aR,6S,8aR)-6-[[(2S)-2-carboxy-4,4-difluoro-1-pyrrolidinyl]-methyl]decahydro-3-isoquinolinecarboxylic acid)], the most potent GLUK5 antagonist described to date. Comparisons were made to the competitive GLUK5/α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor antagonist LY293558 [(3S,4aR,6R,8aR)-6-[2-(1(2)H-tetrazole-5-yl)ethyl]-decahydroisoquinoline-3-carboxylic acid], other decahydroisoquinoline GLUK5 receptor antagonists, and the noncompetitive AMPA receptor antagonist LY300168 [1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodi-azepine]. When characterized electrophysiologically in rat dorsal root ganglion neurons, LY466195 antagonized kainate (30 μM)-induced currents with an IC50 value of 0.045 ± 0.011 μM. In HEK293 cells transfected with GLUK5, GLUK2/GLUK5, or GLUK5/GLUK6 receptors, LY466195 produced IC50 values of 0.08 ± 0.02, 0.34 ± 0.17, and 0.07 ± 0.02 μM, respectively. LY466195 was efficacious in a dural plasma protein extravasation (PPE) model of migraine with an ID100 value of 100 μg/kg i.v. LY466195 was also efficacious in the c-fos migraine model, with a dose of 1 μg/kg i.v. significantly reducing the number of Fos-positive cells in the rat nucleus caudalis after electrical stimulation of the trigeminal ganglion. Furthermore, LY466195 showed no contractile activity in the rabbit saphenous vein in vitro. The diethyl ester prodrug of LY466195 was also efficacious in the same PPE and c-fos models after oral administration at doses of 10 and 100 μg/kg, respectively while having no N-methyl-d-aspartate antagonist-like behavioral effects at oral doses up to 100 mg/kg.

Vasoactive intestinal polypeptide: increased tone, enhancement of acetylcholine release, and stimulation of adenylate cyclase in intestinal smooth muscle.

Abstract Vasoactive intestinal polypeptide (VIP) was examined in vitro for effects on tone and neuronal release mechanisms in intestinal smooth muscle since this is a site of high peptide concentration. VIP contracted the guinea pig ileum and rabbit jejunum in concentrations ranging from 10 −9 to 10 −7 M. Increased tone in the guinea pig ileum was partially antagonized by the anticholinergic agent, atropine (4.38 × 10 −6 M) suggesting that one component of the contractile response was due to the indirect release of acetylcholine. The H 1 receptor antagonist, pyrilamine, did not alter the increased tone produced by VIP indicating that histamine release did not contribute to the ileal contractile response and that VIP exerted a selective effect to enhance neuronal release of acetylcholine. The ability of VIP to modulate acetylcholine release was confirmed in field stimulated ileal preparations where VIP increased the force developed to endogenously released acetylcholine without altering the direct response to acetylcholine. In rabbit jejunum and ileal smooth muscle, VIP related cyclic AMP levels. However, inhibition of phosphodiesterase with papaverine did not potentiate either the VIP-induced ileal contraction or enhancement of the field stimulated response. This raises the possibility that increases in intestinal cyclic AMP may be involved more in VIP-induced alterations in ion transport or secretory phenomenon than in intestinal motility. These studies describing the ability of VIP to modulate acetylcholine release and to increase ileal tone are consistent with the proposed role of VIP in intestinal patholgies involving excessive mucous secretion and motility.

Antagonism of a peripheral vascular but not an apparently central serotonergic response by xylamidine and BW 501C67

Xylamidine and BW501C67 (alpha-anilino-N-2-m-chlorophenoxypropylacetamidine), serotonin antagonists that have been reported not to cross the blood-brain barrier, were compared to other serotonin antagonists: mianserin, ketanserin, metergoline and LY 53857. All six compounds were potent inhibitors of the binding of tritiated spiperone to 5HT2 receptors in rat frontal cortex membranes in vitro and were less potent inhibitors of the binding of tritiated serotonin to 5HT1 receptors in rat brain membranes. All were potent antagonists of the 5HT2 receptor-mediated contractile response to serotonin in the rat jugular vein in vitro. At doses of 0.1 and 0.3 mg/kg i.p., xylamidine and BW501C67 antagonized the pressor response to intravenously injected serotonin in pithed rats. In contrast, neither xylamidine nor BW501C67 at doses of 1 or 3 mg/kg i.p. antagonized the elevation of serum corticosterone concentration by quipazine in rats, although the other compounds antagonized this effect with ED50 values between 0.03 and 0.9 mg/kg. These data corroborate the previously reported antiserotonin activity of xylamidine and BW501C67. Since xylamidine and BW501C67 were potent antagonists of a peripheral serotonergic response in vivo, their inability to antagonize the elevation of serum corticosterone concentration by quipazine suggests that this effect results from activation of central serotonin receptors.

Antagonism of vascular serotonin receptors by m-chlorophenylpiperazine and m-trifluoromethylphenylpiperazine

m-Chlorophenylpiperazine and m-trifluoromethylphenylpiperazine, two compounds that act as agonists at central serotonin receptors mediating certain neuroendocrine, behavioral and serotonin turnover effects, lacked appreciable agonist activity at serotonin receptors mediating contraction of the rat jugular vein. Instead, these compounds were potent antagonists of serotonin-induced contraction of the jugular vein. Apparently these non-indole compounds affect serotonin receptors in various tissues differently, being agonists in brain but mainly antagonists at some peripheral serotonin receptor sites.

Effect of bombesin, bradykinin, substance P and CGRP in prostate, bladder body and neck

Lower urinary tract tissues respond heterogeneously to adrenergic and cholinergic agents. However, the action of bioactive peptides on these tissues has not been extensively studied. The contractile and relaxant effects of nine peptides-bradykinin, cholecystokinin, vasoactive intestinal polypeptide, gastrin, substance P, bombesin, neuropeptide Y, calcitonin gene-related peptide, and motilin-have been compared in the rat bladder body, bladder neck, and left ventral prostate in vitro. All three tissues contracted to bombesin and to bradykinin, although the bladder neck was less sensitive to the contractile effects of bradykinin than the other two tissues. Substance P only contracted the bladder body. Of all the peptides tested, relaxation was only observed to calcitonin gene-related peptide, which relaxed the bladder neck and prostate (phenylephrine-contracted) but not the bladder body (carbamylcholine-contracted). Thus lower urinary tract tissues are responsive to certain bioactive peptides in a nonhomogeneous fashion. These studies raise the possibility that selective modulation of peptide function may be an approach to therapy of urogenital disorders.

Serotonin receptor activation of phosphoinositide turnover in uterine, fundal, vascular, and tracheal smooth muscle.

Hydrolysis of phosphatidylinositul is becoming recognized as the second messenger system for a number of hormones and neurotransmitters, including serotonin. The present study was designed to explore the effects of serotonin to enhance phosphoinositide turnover in several smooth muscle preparations, in an effort (a) to determine which smooth muscle preparation might provide a useful system for further study of phosphoinositide turnover and (b) to examine the role of 5-HT2 receptors in such responses. Basal-[3H] IP(inositol monophosphate ([3H]IP) formation was 10-fold higher in the uterus than in the rat jugular vein, aorta, stomach fundus, or guinea pig trachea. Serotonin produced significant elevations in [3H]IP in the rat aorta, uterus, and jugular vein. Maximal elevation in pH IP was greatest in the jugular vein (eightfold increase) with an ED50 for serotonin of 0.4 (xM. Serotonin (10-7-10−4 M),although a potent contractile agonist in both the guinea pig trachea and rat stomach fundus, did not increase levels in these tissues. The selective 5-HT2 receptor blocker LY53857 (105 M) antagonized the increase in [3H]IP produced by serotonin in the jugular vein, aorta, and uterus. Pargyline (10−6M) added to the trachea and fundus did not unmask an effect of serotonin or enhance the response to serotonin in the rat aorta. Thus, the jugular vein was the tissue most sensitive to activation of pH]IP by serotonin. Increases in PH]IP produced by serotonin may be linked to activation of 5-HT2 receptors in the jugular vein, aorta, and uterus since the selective 5-HT2 antagonist LY53857 could block this biochemical effect. Furthermore, since the guinea pig trachea, like the rat jugular vein, aorta, and uterus, possesses 5-HT: receptors that mediate the contractile response to serotonin and since serotonin did not significantly elevate [3H]IP in the guinea pig trachea, we conclude that activation of 5-HT, receptors may not always result in a measurable increase in [3H]IP.