Mesenchymal stem cell-derived conditioned medium and Methysergide give rise to crosstalk inhibition of 5-HT2A and 5-HT7 receptors in neuroblastoma cells.

OBJECTIVE: (s): We aimed to investigate the effects of mesenchymal stem cell secretome and methysergide combination on 5-hydroxytryptamine 2A, (5-HT2AR), 5-hydroxytryptamine 7 (5-HT7R), adenosine 2A (A2AR) receptors and CD73 on neuroblastoma cell line and how they affect biological characteristics. Methysergide was used as a serotonin antagonist on the neuroblastoma cells. MATERIALS AND METHODS: Human dental pulp-derived stem cells (hDPSCs) used to obtain conditioned medium (CM). Methysergide drug was prepared in CM and applied to neuroblastoma cells. Analysis of 5-HT7R, 5-HT2AR, A2AR and CD73 expressions was performed by western blot and immunofluorescence staining. Total apoptosis, mitochondrial membrane depolarization, Ki-67 proliferation test, viability analysis, DNA damage and cell cycle analysis were performed in accordance with the product procedure by using biological activity test kits. RESULTS: Our results showed that neuroblastoma cancer cells are normally on the Gs signaling axis via the serotonin 7 receptor and the adenosine 2A receptor. CM and Methysergide inhibited the 5-HT7 and A2A receptor levels in neuroblastoma cells. We found that CM and methysergide formed crosstalk inhibition between 5-HT2AR, 5-HT7R, A2AR and CD73. CM and Methysergide increased the total apoptosis in neuroblastoma cells and induced the mitochondrial membrane depolarization. CM and Methysergide induced the DNA damage and arrested in G0/G1 phase of cell cycle of the neuroblastoma cells. CONCLUSION: These findings suggest that the combination of CM and methysergite may exert a therapeutic effect on neuroblastoma cancer cells, and future in vivo studies may be important in area of neuroblastoma research to support the findings.

Involvement of V1-type vasopressin receptors on NaCl intake by hyperosmotic rats treated with muscimol in the lateral parabrachial nucleus.

GABAA receptor activation with agonist muscimol in the lateral parabrachial nucleus (LPBN) induces 0.3 M NaCl intake. In the present study, we investigated water and 0.3 M NaCl intake in male adult rats treated with losartan (angiotensin AT1 receptor antagonist) or MeT-AVP (V1-type vasopressin receptor antagonist) combined with muscimol or methysergide (5-HT2 antagonist) into the LPBN in rats treated with intragastric 2 M NaCl. After 2 M NaCl load and bilateral injections of muscimol (0.5 nmol/0.2 µL) into the LPBN, rats ingested water and 0.3 M NaCl. The pre-treatment of the LPBN with MeT-AVP (1 nmol/0.2 µL) but not losartan (50 µg/0.2 µL) in muscimol treated rats reduced 0.3 M NaCl intake. The pre-treatment of the LPBN with MeT-AVP did not modify the increased 0.3 M NaCl intake in rats treated with methysergide (4 µg/0.2 µL), suggesting that the effect of MeT-AVP was not due to non-specific inhibition of ingestive behavior. The results suggest that endogenous vasopressin in the LPBN facilitates the effects of GABAergic activation driving cell-dehydrated male rats to ingest 0.3 M NaCl.

Involvement of serotonergic and opioidergic systems in the antinociceptive effect of ketamine-magnesium sulphate combination in formalin test in rats.

BACKGROUND: Ketamine and magnesium sulphate showed synergic interaction in the tail-immersion test and additive interaction in the rat formalin test. Aim of study was to evaluate the influence of serotonergic and opioidergic system of this combination in the formalin test in rats. METHODS: Antinociceptive activity was assessed by the formalin test in male Wistar rats (200-250 g). Antagonists (naloxone and methysergide) were administrated 5 min before and magnesium sulphate 5 min after ketamine injection. Formalin (2.5%, 100 µL) was injected into the right hind paw surface (intraplantar) of rats 5 min after ketamine/magnesium combination. Data were recorded as the total time spent in pain related behavior after the injection of formalin or vehicle (0.9% NaCl). RESULTS: In the intermediate phase of the formalin test, methysergide at a dose of 0.2 mg/kg did not have any effect, but at doses of 0.5 and 1 mg/kg it had a pronociceptive effect. Methysergide (0.2, 0.5 and 1 mg/kg) inhibited the antinociceptive effect of ketamine-magnesium sulphate combination. In the intermediate phase, naloxone at a dose of 0.2 mg/kg did not have any effect, but at a dose of 3 mg/kg it produced a pronociceptive effect. Naloxone (0.2 and 3 mg/kg) antagonized the antinociceptive effect of the ketamine (5 mg/kg)-magnesium sulphate (5 mg/kg) combination. CONCLUSION: The results of the present study suggest that serotonergic and opioidergic systems are involved, at least in part, in the antinociceptive effect of the ketamine-magnesium sulphate combination in the model of inflammatory pain in rats.

The antinociceptive effect of anterior pretectal nucleus stimulation is mediated by distinct neurotransmitter mechanisms in descending pain pathways.

Electrical stimulation of the anterior pretectal nucleus (APtN) activates two descending pain inhibitory pathways. One of these pathways relays in the ipsilateral lateral paragigantocellular nucleus (LPGi), whereas the other pathway relays in the contralateral pedunculopontine tegmental nucleus (PPTg). Antinociceptive effect of APtN stimulation has been seen in various pain models in the rodents. Similarly, LPGi or PPTg stimulation results in higher pain thresholds. Descending antinociceptive pathways activated by electrical APtN stimulation have been elucidated, but the underlying neurotransmitter mechanisms involved have not been clarified yet. This study investigates the role that endogenous signaling plays in the ipsilateral LPGi or contralateral PPTg after the APtN is stimulated in the tail-flick test. First, we submitted rats to excitotoxic injection of N-methyl-d-aspartate (NMDA) into the contralateral PPTg. Then, we examined whether blockage of NMDA (AP-7), serotonergic (methysergide), or opioid (naloxone) receptors in the ipsilateral LPGi is required for APtN stimulation-evoked analgesia (SEA). Likewise, we examined the effects of antagonists of NMDA, serotonergic, or cholinergic nicotinic (mecamylamine) receptors on the contralateral PPTg in ipsilateral LPGi-lesioned rats. Our results confirmed that APtN stimulation activates two pain inhibitory pathways and showed that endogenous opioid signaling in the ipsilateral LPGi appears to be necessary for APtN SEA and for endogenous NMDA, serotoninergic, and nicotinergic signaling in the contralateral PPTg.

The effect of serotonin on penicillin-induced epileptiform activity.

OBJECTIVE: This study was aimed at examining the epileptiform activity of the 5-HT2 serotonin receptor agonist and antagonist, and 5-hydroxytryptophan (5-HTP) in penicillin-induced epilepsy in albino Wistar rats. METHODS: For this purpose, 90 albino male Wistar rats were used in this study. Epileptiform activity was induced by an injection of penicillin, an agonist of GABAA receptor, (500 IU, i.c.) into the somatomotor cortex. Thirty minutes after the injection of penicillin, 2,5-dimethoxy-4-iodoamphetamine (DOI, an agonist of 5-HT2 receptor) (0.5, 1, 2 and 4 mg/kg, i.p.), methysergide, an antagonist of 5-HT2 receptor, (1, 10, 20, 50 and 100 µM, i.c.v.) and 5-HTP, precursor of 5-HT, (25, 50, 75 and 100 mg/kg, i.p.) were administered, respectively. RESULTS: DOI, at the doses of 1 and 2 mg/kg, significantly decreased penicillin-induced epileptiform activity (p < 0.05). Methysergide, at the doses of 20, 50 and 100 µM, significantly increased the mean spike frequency of penicillin-induced epileptiform activity (p < 0.05). The doses of 50, 75 and 100 mg/kg of 5-HTP decreased the mean spike frequency of penicillin-induced epileptiform activity (p < 0.05). The mean of amplitude of penicillin-induced epileptiform activity did not significantly change in any of the groups (p > 0.05). CONCLUSION: The electrophysiological data from the present study suggest that serotonin 5-HT2 receptors have an important role in controlling penicillin-induced epileptiform activity in the rat.

Breathing stimulation mediated by 5-HT1A and 5-HT3 receptors within the preBötzinger complex of the adult rabbit.

Serotonin (5-HT) has been reported to play excitatory effects on respiration by acting on preBötzinger complex (preBötC) neurons in neonatal or juvenile rodents. However, whether its action is circumscribed to the preBötC and present in other animal species, particularly in adult preparations, is unknown. We investigated the respiratory role of 5-HT within the preBötC and neighbouring respiration-related regions. Experiments were performed on α-chloralose-urethane anesthetized, vagotomized, paralyzed and artificially ventilated rabbits making use of bilateral microinjections (30-50 nl). 5-HT caused excitatory effects on respiratory activity only when applied to the preBötC. These effects were mediated by 5-HT1A and 5-HT3 receptors as shown by microinjections of specific agonists of the different types of 5-HT receptors. Unexpectedly, the blockade of 5-HT1A receptors by methysergide or the specific antagonist (S)-WAY 100135 induced excitatory respiratory effects. Microinjections of the 5-HT3 receptor antagonist ondansetron did not influence respiration, but prevented (S)-WAY 100135-induced responses. The blockade of GABAA receptors by bicuculline within the preBötC prevented the effects of the 5-HT1A receptor agonist 8-OH-DPAT. The involvement of GABAergic inhibition and 5-HT1A receptor-mediated disinhibition is also corroborated by immunohistochemical data. The results show for the first time in an adult animal preparation that 5-HT plays a pivotal role in the modulation of the preBötC activity probably via both presynaptic and postsynaptic mechanisms and highlight the importance of disinhibition phenomena. Present findings may be relevant to some respiratory disorders in which an impairment of central 5-HT mechanisms has been reported, such as sleep apnoea and sudden infant death syndrome.

Phrenic long-term depression evoked by intermittent hypercapnia is modulated by serotonergic and adrenergic receptors in raphe nuclei.

Intermittent hypercapnia evokes prolonged depression of phrenic nerve activity (phrenic long-term depression, pLTD). This study was undertaken to investigate the role of 5-HT and α2-adrenergic receptors in the initiation of pLTD. Adult male urethane-anesthetized, vagotomized, paralyzed, and mechanically ventilated Sprague-Dawley rats were exposed to a protocol of acute intermittent hypercapnia (AIHc; 5 episodes of 15% CO2 in air, each episode lasting 3 min). The experimental group received microinjection of the selective 5-HT1A receptor agonist 8-hydroxy-2-(dipropylamino)tetralin hydrobromide (8-OH-DPAT), the broad-spectrum 5-HT antagonist methysergide, or the α2-adrenergic antagonist yohimbine, whereas the control group received microinjection of 0.9% saline into the caudal raphe region. Peak phrenic nerve activity (pPNA) and burst frequency ( f) were analyzed during baseline (T0), during 5 hypercapnic episodes (THc1-THc5), and at 15, 30, and 60 min after the end of the last hypercapnic episode. In the control group, pPNA decreased 60 min after the end of the last hypercapnic episode compared with baseline values, i.e., pLTD developed ( P = 0.023). In the 8-OH-DPAT group, pPNA significantly decreased at T15, T30, and T60 compared with baseline values, i.e., pLTD developed ( P = 0.01). In the methysergide and yohimbine groups, AIHc did not evoke significant changes of the pPNA at T15, T30, and T60 compared with baseline values. In conclusion, activation of 5-HT1A receptors accentuated induction of pLTD, whereas blockade of α2-adrenergic receptors prevented development of pLTD following AIHc in anesthetized rats. These results suggest that chemical modulation of 5-HT and α2-adrenergic receptors in raphe nuclei affects hypercapnia-induced pLTD, offering important insights in understanding the mechanisms involved in development of respiratory plasticity. NEW & NOTEWORTHY Hypercapnia is a concomitant feature of many breathing disorders, including obstructive sleep apnea. In this study, acute intermittent hypercapnia evoked development of phrenic long-term depression (pLTD) 60 min after the last hypercapnic episode that was preserved if the selective 5-HT1A receptor agonist 8-hydroxy-2-(dipropylamino)tetralin hydrobromide was microinjected in the caudal raphe region before the hypercapnic stimulus. This study highlights that both 5-HT and adrenergic receptor activation is needed for induction of pLTD in urethane-anesthetized rats following intermittent hypercapnia exposure.

Presynaptic serotonin 5-HT1B/D receptor-mediated inhibition of glycinergic transmission to the frog spinal motoneurons.

Endogenous monoamine 5-hydroxytryptamine (5-HT, serotonin) is a phylogenetically ancient neurotransmitter present in vertebrates. The functions of 5-HT in central nervous system are intensively studied; however, the presynaptic effects of 5-HT in frog spinal motoneurons are practically unexplored. We have previously shown that 5-HT decreases the frequency of glycinergic miniature inhibitory postsynaptic potentials (mIPSPs), but does not affect the frequency of GABAergic mIPSPs and increases the frequency of glutamatergic postsynaptic potentials. In the present study, using pharmacological methods and intracellular recordings in motoneurons from an adult frog's isolated spinal cord, we aimed to identify the 5-HT receptor subtype responsible for inhibiting the release of glycine. Аn agonist of 5-HT1A and 5-HT7 receptors, 8-OH-DPAT, and a selective agonist of 5-HT2 receptors, α-Ме-5-НТ, did not show any significant effect on inhibitory transmission, indicating that 5-HT1A, 5-HT2, and 5-HT7 receptors are not involved in the modulation of glycine release in the adult frog spinal cord. An agonist of 5-HT1B/D receptors sumatriptan decreased the frequency (but not the amplitude) of glycinergic mIPSPs similar to 5-HT. An antagonist of 5-HT1,2 receptors, methysergide, abolished the effect of sumatriptan. Together our results suggest that 5-HT inhibits the release of glycine by activation of 5-HT1B/D receptors.

Daily acute intermittent hypoxia improves breathing function with acute and chronic spinal injury via distinct mechanisms.

Daily acute intermittent hypoxia (dAIH) elicits respiratory plasticity, enhancing respiratory motor output and restoring breathing capacity after incomplete cervical spinal injuries (cSCI). We hypothesized that dAIH-induced functional recovery of breathing capacity would occur after both acute (2 weeks) and chronic (8 weeks) cSCI, but through distinct cellular mechanisms. Specifically, we hypothesized that dAIH-induced breathing recovery would occur through serotonin-independent mechanisms 2wks post C2 cervical hemisection (C2Hs), versus serotonin-dependent mechanisms 8wks post C2Hs. In two independent studies, dAIH or sham (normoxia) was initiated 1 week (Study 1) or 7 weeks (Study 2) post-C2Hs to test our hypothesis. Rats were pre-treated with intra-peritoneal vehicle or methysergide, a broad-spectrum serotonin receptor antagonist, to determine the role of serotonin signaling in dAIH-induced functional recovery. Our data support the hypothesis that dAIH-induced recovery of breathing capacity transitions from a serotonin-independent mechanism with acute C2Hs to a serotonin-dependent mechanism with chronic C2Hs. An understanding of shifting mechanisms giving rise to dAIH-induced respiratory motor plasticity is vital for clinical translation of dAIH as a therapeutic modality.

Antitussive and expectorant activities of licorice and its major compounds.

Licorice has been used as an antitussive and expectorant herbal medicine for a long history. This work evaluated the activities of 14 major compounds and crude extracts of licorice, using the classical ammonia-induced cough model and phenol red secretion model in mice. Liquiritin apioside (1), liquiritin (2), and liquiritigenin (3) at 50 mg/kg (i.g.) could significantly decrease cough frequency by 30-78% (p < .01). The antitussive effects could be partially antagonized by the pretreatment of methysergide or glibenclamide, but not naloxone. Moreover, compounds 1-3 showed potent expectorant activities after 3 days treatment (p < .05). The water and ethanol extracts of licorice, which contain abundant 1 and 2, could decrease cough frequency at 200 mg/kg by 25-59% (p < .05), and enhance the phenol red secretion (p < .05), while the ethyl acetate extract showed little effect. These results indicate liquiritin apioside and liquiritin are the major antitussive and expectorant compounds of licorice. Their antitussive effects depend on both peripheral and central mechanisms.

Participation of Opioid Pathway in the Central Antinociceptive Effects of Eugenol

The aim of the present study was to evaluate the central antinociceptive effects of eugenol after intraperitoneal administration. Experiments were carried out using male Sprague-Dawley rats. Subcutaneous injection of 5% formalin-induced nociceptive behavioral responses was used as the pain model. Subcutaneous injection of 5% formalin significantly produced nociceptive responses by increasing the licking time during nociceptive behavior. Subsequent intraperitoneal injection of 100 mg/kg of eugenol led to a significant decrease in the licking time. However, low dose of eugenol (50 mg/kg) did not affect the nociceptive behavioral responses produced by subcutaneous injection of formalin. Intrathecal injection of 30 µg of naloxone, an opioid receptor antagonist, significantly blocked antinociceptive effects produced by intraperitoneal injection of eugenol. Neither intrathecal injection of methysergide (30 µg), a serotonin receptor antagonist nor phentolamine (30 µg), an α-adrenergic receptor antagonist influenced antinociceptive effects of eugenol, as compared to the vehicle treatment. These results suggest that central opioid pathway participates in mediating the antinociceptive effects of eugenol.

Antinociceptive effects of low-level laser therapy at 3 and 8 j/cm2 in a rat model of postoperative pain: possible role of endogenous Opioids.

Low-level laser therapy (LLLT) is the direct application of light to stimulate cell responses (photobiomodulation) to promote tissue healing, reduce inflammation, and induce analgesia; the molecular basis for these effects of LLLT remains unclear. The objective of this study was to evaluate the analgesic effect of LLLT in the rat plantar incision model of postoperative pain as well as to investigate some of the possible mechanisms involved in this effect. Wistar rats were submitted to plantar incision and treated with LLLT (830 nm, continuous-mode, 30 mW/cm2 , 1-12 J/cm2 ). Postoperative thermal and mechanical hypersensitivity were monitored for 24 hours post-incision. In addition, the animals were pretreated with saline, naloxone (a nonselective opioid receptor antagonist; 20 µg/5 µl) or methysergide (5-HT2C , 5-HT2A , 5-HT7 , 5-HT5a , 5-HT6, and 5-HT1F receptors antagonist; 30 µg/5 µl). Moreover, 24 hours after incision and treatment, the TNF-α and IL-1ß levels in serum were evaluated. Our results demonstrate, for the first time, that LLLT at 3 or 8 J/cm2 , but not at 1-2, 4-7, or 9-12 J/cm2 , induced an analgesic effect on postoperative pain. Naloxone, but not methysergide, blocked the LLLT-induced anti-nociceptive effect. Additionally, IL-1-ß and TNF-α production significantly decreased after LLLT at 3 or 8 J/cm2 . Our results suggest that LLLT at 3 or 8 J/cm2 primarily modulates the endogenous opioids system and is not directly mediated by serotonergic receptors. Reduction of IL-1ß and TNF-α may play a role in the antinociceptive action of LLLT. Lasers Surg. Med. 49:844-851, 2017. © 2017 Wiley Periodicals, Inc.

Anti-hyperalgesic effect of Lippia grata leaf essential oil complexed with ß-cyclodextrin in a chronic musculoskeletal pain animal model: Complemented with a molecular docking and antioxidant screening.

BACKGROUND: Due to its unclear pathophysiology, the pharmacological treatment of fibromyalgia is a challenge for researchers. Studies using medicinal plants, such as those from the genus Lippia, complexed with cyclodextrins (CDs) have shown innovative results. OBJECTIVE: The present research intended to evaluate the effect of an inclusion complex containing ß-cyclodextrin (ßCD) inclusion complex with Lippia grata (LG) essential oil in a chronic musculoskeletal pain model, its central activity and its possible interaction with neurotransmitters involved in pain. METHODS: After acid saline-induced chronic muscle pain, male mice were evaluated for primary and secondary hyperalgesia and muscle strength. Moreover, an antagonist assay was performed to assess the possible involvement of the opioidergic, serotonergic and noradrenergic pathways. In addition, Fos protein in the spinal cord was assessed, and a docking study and antioxidant assays were performed. RESULTS: The treatment with LG-ßCD, especially in the dose of 24mg/kg, was able to significantly decrease (p<0.05) the paw withdrawal and muscle threshold. Furthermore, LG-ßCD was shown to affect the opioidergic and serotonergic pathways. There were no significant changes in muscle strength. Fos protein immunofluorescence showed a significant decrease in expression in the dorsal horn of the spinal cord. The main compounds of LG showed through the docking study interaction energies with the alpha-adrenergic and µOpioid receptors. In all antioxidant assays, LG exhibited stronger antioxidant activities than LG-ßCD. CONCLUSION: This study suggested that LG-ßCD could be considered as a valuable source for designing new drugs in the treatment of chronic pain, especially musculoskeletal pain.

Effects of the Novel High-affinity 5-HT(1B/1D)-receptor Ligand Frovatriptan on the Rat Carotid Artery.

BACKGROUND: In blood vessels 5-HT stimulates sympathetic nerves, the endothelium and vascular smooth muscle cells. Triptans are specific anti-migraine drugs and they activate the serotoninergic 5HT1b/d receptors causing vasoconstriction of the cerebral vessels. AIM: To evaluate the effect of frovatriptan on isolated rat carotid artery. METHODS: Contractile activity of the preparations was registered isometrically. Krebs solution (pH = 7.4) was used for washing smooth muscle (SM) preparations aerated with 95% O2 and 5% CO2 at 37°C. The 60-minute adaptation of tone level of preparations was taken as a starting tone and the changes such as contraction or relaxation were calculated using it. RESULTS: Frovatriptan (1×10-6 mol/l - 1×10-5 mol/l) induced a contraction, but at higher concentrations it caused relaxation of the carotid artery. The L-norepinephrine contractile reaction was enhanced in the presence of frovatriptan. In the presence of 5-HT2 receptor antagonist, methysergide, frovatriptan increased the relaxation. In the presence of the specific α-1 receptor antagonist, prazosin, the frovatriptan-induced relaxation decreased. CONCLUSION: The observed contractile effect of frovatriptan is probably associated with the main effect of the drug - activation of the serotoninergic 5HT1B /1D receptors causing vasoconstriction of the cerebral vessels and their anti-migraine effect. At higher concentrations, frovatriptan, most likely via some non-specific mechanism, could activate the following intracellular chain reaction: stimulation of α1D could activate eNOS which may increase in the concentration of NO which results in the final effect of relaxation.

The role of methysergide in migraine and cluster headache treatment worldwide - A survey in members of the International Headache Society.

Background Methysergide has been as an effective treatment for migraine and cluster headache for over 50 years but has recently been investigated by the European Medicines Agency due to safety concerns. Methods To assess the need for continuing availability of methysergide, the International Headache Society performed an electronic survey among their members. Results The survey revealed that 71.3% of all respondents had ever prescribed methysergide and 79.8% would prescribe it if it were to become available. Respondents used it more in cluster headache than migraine, and reserved it for use in refractory patients. Conclusion The vast majority of headache experts in this survey regarded methysergide a unique treatment option for specific populations for which there are no alternatives, with an urgent need to continue its availability. This position was supported by the International Headache Society.

Drug-induced aortic valve stenosis: An under recognized entity.

BACKGROUND: We have been intrigued by the observation that aortic stenosis (AS) may be associated with characteristic features of mitral drug-induced valvular heart disease (DI-VHD) in patients exposed to valvulopathic drugs, thus suggesting that beyond restrictive heart valve regurgitation, valvulopathic drugs may be involved in the pathogenesis of AS. METHODS: Herein are reported echocardiographic features, and pathological findings encountered in a series of patients suffering from both AS (mean gradient >15mmHg) and mitral DI-VHD after valvulopathic drugs exposure. History of rheumatic fever, chest radiation therapy, systemic disease or bicuspid aortic valve disease were exclusion criteria. RESULTS: Twenty-five (19 females, mean age 62years) patients having both AS and typical features of mitral DI-VHD were identified. Mean transaortic pressure gradient was 32+/-13mmHg. Aortic regurgitation was ≥ mild in 24 (96%) but trivial in one. Known history of aortic valve regurgitation following drug initiation prior the development of AS was previously diagnosed in 17 patients (68%). Six patients underwent aortic valve replacement and 3 both aortic and mitral valve replacement. In the 9 patients with pathology analysis, aortic valvular endocardium was markedly thickened by dense non-inflammatory fibrosis, a characteristic feature of DI-VHD. CONCLUSION: The association between AS and typical mitral DI-VHD after valvulopathic drug exposure may not be fortuitous. Aortic regurgitation was usually associated to AS and preceded AS in most cases but may be lacking. Pathology demonstrated the potential role of valvulopathic drugs in the development of AS.

Anxiolytic effect of the GPR103 receptor agonist peptide P550 (homolog of neuropeptide 26RFa) in mice. Involvement of neurotransmitters.

The GPR103 receptor is a G protein-coupled receptor, which plays a role in several physiological functions. However, the role of the GPR103 receptor in anxiety has not been clarified. The first aim of our study was to elucidate the involvement of the GPR103 receptor in anxious behavior. Mice were treated with peptide P550, which is the mouse homolog of neuropeptide 26RFa and has similar activity for the GPR103 receptor as neuropeptide 26RFa. The anxious behavior was investigated using an elevated plus-maze paradigm. The second aim of our study was to investigate the underlying neurotransmissions. Accordingly, mice were pretreated with a nonselective muscarinic acetylcholine receptor antagonist, atropine, a γ-aminobutyric acid subunit A (GABAA) receptor antagonist, bicuculline, a non-selective 5-HT2 serotonergic receptor antagonist, cyproheptadine, a mixed 5-HT1/5-HT2 serotonergic receptor antagonist, methysergide, a D2, D3, D4 dopamine receptor antagonist, haloperidol, a nonselective α-adrenergic receptor antagonist, phenoxybenzamine and a nonselective ß-adrenergic receptor antagonist, propranolol. Our results demonstrated that peptide P550 reduces anxious behavior in elevated plus maze test in mice. Our study shows also that GABAA-ergic, α- and ß-adrenergic transmissions are all involved in this action, whereas 5-HT1 and 5-HT2 serotonergic, muscarinic cholinergic and D2, D3, D4 dopaminergic mechanisms may not be implicated.

Effects of cyclopiazonic acid and dexamethasone on serotonin-induced calcium responses in vascular smooth muscle cells.

We previously observed that sarcoendoplasmic reticulum Ca(2+) ATPase (SERCA) blockade by cyclopiazonic acid (CPA) significantly potentiates serotonin (5-hydroxytryptamine (5-HT))-induced vascular contractions. Furthermore, 5-HT receptor antagonist methysergide partially inhibited CPA-potentiated 5-HT contractions. In the present study, we further investigated whether SERCA inhibition potentiates 5-HT-induced Ca(2+) responses along with attenuating the receptor antagonism by store-operated Ca(2+) (SOC) entry and protein kinase C (PKC)-mediated mechanisms. The effects of dexamethasone that was previously shown to induce SOC entry and enhance 5-HT responses were also tested. For this purpose, intracellular Ca(2+) levels were monitored in A7r5 embryonic rat vascular smooth muscle cells by spectrofluorometry using the fluorescent indicator fura-2. The results showed that CPA, although not dexamethasone, significantly potentiated 5-HT-induced Ca(2+) elevations. Ketanserin partially decreased 5-HT-induced and CPA-potentiated Ca(2+) elevations whereas both PKC inhibitor D-sphingosine and SOC entry blocker 2-aminoethoxydiphenyl borate (2-APB) abolished the remaining responses. The data suggests that diminished antagonistic effect on 5-HT-induced Ca(2+) elevations in the presence of SERCA inhibition is induced by SOC entry and PKC activation.

Intermittent hypercapnia-induced phrenic long-term depression is revealed after serotonin receptor blockade with methysergide in anaesthetized rats.

NEW FINDINGS: What is the central question of this study? Intermittent hypercapnia is a concomitant feature of breathing disorders. Hypercapnic stimuli evoke a form of respiratory plasticity known as phrenic long-term depression in experimental animals. This study was performed to investigate the putative role of serotonin receptors in the initiation of phrenic long-term depression in anaesthetized rats. What is the main finding and its importance? Phrenic nerve long-term depression was revealed in animals pretreated with the serotonin broad-spectrum antagonist, methysergide. This study highlights that serotonin receptors modulate respiratory plasticity evoked by acute intermittent hypercapnia in anaesthetized rats. This study was performed to test the hypothesis that intermittent hypercapnia can evoke a form of respiratory plasticity known as long-term depression of the phrenic nerve (pLTD) and that 5-HT receptors play a role in the initiation of pLTD. Adult male urethane-anaesthetized, vagotomized, paralysed, mechanically ventilated Sprague-Dawley rats were exposed to an acute intermittent hypercapnia protocol. One group received i.v. injection of the non-selective 5-HT receptor antagonist methysergide and another group received i.v. injection of the selective 5-HT1A receptor antagonist WAY-100635 20 min before exposure to intermittent hypercapnia. A control group received i.v. injection of saline. Peak phrenic nerve activity and respiratory rhythm parameters were analysed at baseline (T0), during each of five hypercapnic episodes, and 15, 30 and 60 min (T60) after the last hypercapnia. Intravenous injection of methysergide before exposure to acute intermittent hypercapnia induced development of amplitude pLTD at T60 (decreased by 46.1 ± 6.9%, P = 0.003). Conversely, in control and WAY-100635-pretreated animals, exposure to acute intermittent hypercapnia did not evoke amplitude pLTD. However, a long-term decrease in phrenic nerve frequency was evoked both in control (42 ± 4 breaths min(-1) at T0 versus 32 ± 5 breaths min(-1) at T60; P = 0.036) and in methysergide-pretreated animals (42 ± 2 breaths min(-1) at T0 versus 32 ± 3 breaths min(-1) at T60; P = 0.028). In WAY-100635 pretreated animals, frequency pLTD was prevented. These results suggest that 5-HT receptors modulate respiratory plasticity induced by acute intermittent hypercapnia in anaesthetized rats.

[Cluster headache treatment]. / Traitements de l'algie vasculaire de la face.

UNLABELLED: Acute treatment: sumatriptan, oxygen inhalation. Prophylactic treatment: verapamil, lithium carbonate. Transitional treatment. SURGICAL TREATMENT: deep brain stimulation, occipital nerve stimulation, stimulation of the sphenopalatin ganglion.