Imetit Dihydrobromide and Thioperamide Medication in Cough Hypersensitivity Model—The Role of H3 Receptors

Chronic cough is a troublesome problem and it is frequently associated with diseases such as gastroesophageal reflux, asthma and upper airway diseases—so called diagnostic triade. The magnitude and severity of cough is strongly associated with the ongoing nasal inflammation in subjects with rhinosinusitis and treatment of nasal inflammation leads to the down regulation of pathologically up-regulated cough. Histamine plays a key role in the inflammation of the upper airways of different aetiologies;therefore histamine receptors seem to be promising targets. The aim of our study was to ascertain the effect of H3R agonist imetit and H3R antagonist thioperamide on cough and symptoms of allergic rhinitis (AR) in an animal model of upper airway cough syndrome in ovalbumin sensitized guinea pigs. OVA sensitized guinea pigs (n = 10) were repeatedly challenged with i.n. allergen-OVA to induce allergic rhinitis and to enhance cough reflex according to the validated model of experimental allergic rhinitis. Animals were pre-treated by i.p. administration of imetit (1 mg/kg and 2 mg/kg of body weight) and thioperamide 30 min. prior i.n. OVA administration. Rhinitis evaluation was based on the occurrence of typical symptoms. The effect on cough was assessed from the response to inhalation of citric acid (0.4 M, 10 min), final cough count and cough latency were analysed from the airflow traces, cough motor pattern and the cough sound. AR up-regulated the cough response from 9 ± 2 to 16 ± 1 cough per provocation, med ± IQR, p < 0.05 and shortened cough latency. Imetit (1 mg/kg) suppressed nasal symptoms and decreased number of cough from 16 ± 1 to 12 ± 1;however the data did not reach significance. Imetit (2 mg/kg) significantly suppressed the nasal symptoms, and number of coughs from 16 ± 1 to 6 ± 2, med ± IQR, p < 0.05. Thioperamide (5 mg/kg of body weight) did not have expected effects on tested parameters. H3R agonist imetit, unlike H3R antagonist thioperamide has antitussive potential and ability to suppress nasal symptoms in animal model of allergic rhinitis.

Changes in hippocampal histamine receptors in rats after treatment with Trimeresurus albolabris venom

BACKGROUND： It has been demonstrated that histamine and its receptors in the hippocampus play an important role in memory and/or learning behaviors.OBJECTIVE： To investigate the expression levels of the histamine receptor gene and protein in the hippocampi of rats prior to and after administration of Trimeresurus albolabris venom using reverse transcription-polymerase chain reaction （RT-PCR） and Western blot techniques. DESIGN, TIME AND SETTING： A controlled observation based on cellular protein level was performed in the College of Life Sciences, Chongqing Normal University between March 2005 and April 2007. MATERIALS： Eighty adult male Sprague-Dawley rats were provided by the Laboratory Animal Center of the Third Military Medical University of Chinese PLA. The lyophilized powder of Trimeresurus albolabris venom was collected from Jin-Hu-Shan in Chongqing, China. METHODS： Twenty rats were randomly and evenly divided into an experimental group and a control group The experimental group was subcutaneously injected with 0.65 mg/mL Trimeresurus albolabris venom, 0.5 mL for each rat. The control group was subcutaneously injected with an equal amount of 0.9% physiological saline. Prior to and after injection, rats from these two groups were placed in the Morris Water Maze for recording of path length and escape latency. The remaining 60 rats were randomly allocated to another experimental group （n = 50） and another control group （n = 10）. Rats were correspondingly injected as described above. At different time points （0.1, 0.5, 1, 2, and 3 hours after injection）, rats were decapitated and bilateral hippocampal tissues were dissociated （approximately 100 mg for each sample）. Then, the acquired hippocampal tissue was immediately preserved at -70 ℃ for subsequent experiments. MAIN OUTCOME MEASURES： （1） The levels of histamine receptor （including H1R, H2R, and H3R） mRNA and protein in the hippocampi of rats were measured prior to and after injection of Trimeresurus albolabris venom using RT-PCR and Western Blot techniques. （2） Escape latency （namely, time to reach a platform） and path length were examined by Morris Water Maze testing. RESULTS： All 80 rats were included in the final analysis. In the experimental group, the level of mRNA for H3R receptor in rat hippocampi was just slightly changed, but the level of H3R receptor protein was significantly down-regulated compared with that in the control group （P 〈 0.05）. Both mRNA and protein levels for H1R receptor were initially downregulated and then recovered to normal levels. Expression of H2R receptor mRNA was initially upregulated, then downregulated, and finally restored to the control level. The level of H2R receptor protein showed a tendency for downregulation. In the Morris Water Maze testing, escape latency and path length were significantly longer in the experimental group than in the control group （P 〈 0.05）. CONCLUSION： Within three hours of injection with Trimeresurus albolabris venom, mRNA and protein levels of most histamine receptors in rat hippocampi were downregulated. Such changes possibly contribute to an impairment of memory and/or learning behaviors in rats following injection of Trimeresurus albolabris venom.

Effect of Fructus Psoraleae on motility of gallbladder isolated smooth muscle strips from guinea pigs

AIM： To observe the effect of Fructus Psoraleae on motility of isolated gallbladder muscle strips of guinea pigs and its mechanism. METHODS： Guinea pigs were hit to lose consciousness and the whole gallbladder was removed quickly. Two or three smooth muscle strips （8 mm×3mm） were cut along a longitudinal direction. The mucosa was gently removed. Every longitudinal muscle strip was suspended in a tissue chamber which was continuously perfused with 5 mL Krebs solution （37℃）, pH 7.4, and aerated with 950 mL/L 02 and 50 mL/L CO2. The isometric response was recorded with an ink-writing recorder. After 2 h equilibration under i g-load, 50 μL Fructus Psoraleae （10, 20, 70, 200, 700, 1000 g/L） was added cumulatively into the tissue chamber in turn every 2 rain to observe their effects on gallbladder muscle strips （cumulating final concentration of Fructus Psoraleae was 0.1, 0.3, 1.0, 3.0, 10.0, 20.0 g/L）. The antagonists, including 4-DAMP, benzhydramine, hexamethonium, phentolamine, verapamil and idomethine were given 2 min before Fructus Psoraleae respectively to investigate the mechanisms involved. RESULTS： Fructus Psoraleae dose-dependently increased the resting tension （r=0.992, P〈0.001）, decreased the mean contractile amplitude （r=0.970, P〈0.001） and meanwhile increased the contractile frequency of the gallbladder muscle strip in vitro （r=0.965, P〈0.001）. The exciting action of Fructus Psoraleae on the resting tension could be partially blocked by 4-DAMP （the resting tension decreased from 1.37 ± 0.41 to 0.70 ± 0.35, P〈0.001）, benzhydramine （from 1.37 ±0.41 to 0.45±0.38, P〈0.001）, hexamethonium （from 1.37 ± 0.41 to 0.94 ± 0.23, P〈0.05）, phentolamine （ from 1.37±0.41 to 0.89±0.22, P〈0.01） and verapamil （from 1.37±0.41 to 0.94±0.26, P〈0.05）. But the above antagonists had no significant effect on the action of Fructus Psoraleae-induced mean contractile amplitude （P〉0.05）. Moreover, the increase of the contractile frequency due to Fructus Psoraleae was inhibited by 4-DAMP （decreased from 8.3 ± 1.2 to 6.8 ± 0.5, P 〈 0.01） and hexamethonium （from 8.3 ±1.2 to 7.0 ± 0.9, P 〈 0.05）. Idomethine had no significant effect on the Fructus Psoraleae- induced responses （P〉 0.05）. CONCLUSION： Fructus Psoraleae enhances the motility of isolated gallbladder muscle strips from guinea pigs, in a dose-dependent manner. The effect of Fructus Psoraleae is partly related to M3, N receptor, α receptor, H1 receptor, Ca^2＋ channel, but not related to prostaglandin.

Effects of splice sites on the intron retention in histamine H_3 receptors from rats and mice

In the alternative splicing, intron retention, of histamine H3 receptors in rats and mice, the short transcript isoforms that are excised alternatively spliced introns are easily detected in a very low level in rats and are undetectable in mice using the regular PCR protocol. The retained introns have common 5＇ splice site and different 3＇ splice sites. The detailed mechanism for the special alternative splicing remains largely unclear. In this study, we developed a minigene splicing system to recapitulate natural alternative splicing of the receptors and investigated the effects of 5＇ and 3＇ splice sites on intron retention in HeLa cells. Mutating weak 5＇ and 3＇ splice sites of the alternatively spliced introns toward the canonical consensus sequences promoted the splicing of the corresponding introns in rat and mouse minigenes. The effect of splice site strength was context-dependent and much more sigiaificant for the 3＇ splice site of the longer alternative intron than for the 3＇ splice site of the shorter alternative intron and the common 5＇ splice sites; it was also more significant in the rat minigene than in the mouse minigene. Mutating the 3＇ splice site of the longer alternative intron resulted in almost complete splicing of the intron and made the corresponding isoform to become the nearly exclusive transcript in the rat minigene.