| Objective: The aim of this study was to investigate the alteration ofcontractile and relaxant responses of the isolated gastric smooth muscle inearly diabetic rats, and to analyze the effect and mechanism of berberine onimproving the neural function of diabetic rats’ gastric smooth muscle.Methods: Male SD rats (the body weight at200g) were divided into5groups: normal rats group, diabetic rats group, diabetic rats treated with highor low dose berberine by gavage group and diabetic rats treated with berberineby intraperitoneal injection group. Rats in diabetic group were fasted for12hbefore administration streptozotocin (65mg·kg-1in1%sodium citrate buffer)by intraperitoneal injection. After two weeks of the streptozotocin-administration, the rats in high/low dose gavage group were given berberine(200/100mg·kg-1) by intragastric administration. The rats in intraperitonealinjection group were injected berberine (20mg·kg-1), once a day for two weeks.Using the method of organ bath, contractile and relaxant responses wereinduced by electric field stimulation (EFS) and pharmacology agonist ongastric smooth musclein rats, and the tool drugs was used to intervene in theneurotransmitter activity. We observe the alteration of contractile and relaxantresponses of the gastric smooth muscle in early diabetic rats, and analyze thechanging of cholinergic, nitriergic and purinergic nerve in neurogeniccontractile and relaxant responses. Based on this, we explore the effects ofberberine on the neural function of gastric smooth muscle in diabetic rats bydifferent administration routes, and analyze the possible mechanisms.Results1. Changes of the symptoms and the functions of the gastric fundus smoothmuscle in the early diabetic rats1.1The alterations of the status, the level of blood glucose and gastric morphology in diabetic ratsCompared with normal rats which had the same week-old, the drinkingand urinating activities were increased in the diabetic rats, and the hair and tailcolour were much darker; body weight was reduced (P<0.01), and the level ofblood glucose was increased significantly (P<0.01).The gastric tissues of diabetic rats became much weaker, limper andlarger, especially the changes of gastric fundus; the stomach remains somecontents and appears abnormal phenomenon in gastric emptying in diabeticrats.1.2The functional alterations in the gastric body and gastric antrum smoothmuscle in diabetic ratsThere were no significant differences in the EFS-induced contractions ongastric body smooth muscle and the spontaneous contractions on gastricantrum in normal and diabetic rats (P>0.05).1.3The contractile responses induced by EFS in gastric fundus smooth muscleThe reproducible and stable contractile responses were induced by EFS(20Hz), and could be completely blocked by1μmol·L-1atropine or0.1μmol·L-1TTX on gastric fundus smooth muscle in normal rats; but indiabetic rats, the contractile responses were smaller, and the contractileamplitudes were apparently disordered, compared with that of in normal rats(P<0.05)1.4The relaxant responses induced by EFS in gastric fundus smooth muscleand pylorus circular muscleThe reproducible and frequency-dependent relaxant responses wereinduced by EFS (1-10Hz), and could be completely blocked by0.1μmol·L-1TTX on gastric fundus smooth muscle and pylorus circular muscle in normaland diabetic rats. Compared with normal rats, FES-induced relaxations weresignificantly larger on gastric fundus smooth muscle in diabetic rats (P<0.01),but the contrary results were gotten on pylorus circular muscle (P<0.01).When the strips were pre-contracted by0.3μmol·L-1carbachol,100μmol·L-1L-NAME significantly inhibited EFS-induced relaxations on gastric fundus circular muscle, and no differences between the normal anddiabetic rats (P<0.05), the percentage of inhibition on the relaxation was94.8%and93.7%at1Hz, respectively. The EFS-induced relaxations werefurther inhibited when L-NAME combined with α–chymotrypsin or suramin(P<0.05), and were more effective than that of using L-NAME only.2. Effects of berberine on the functions of gastric fundus and pylorus circularmuscle in diabetic rats and the related mechanisms2.1Effects of berberine on the contractile responses to EFS in gastric funduscircular muscle and mechanisms in virto and in vivo administrationBerberine corrected the disorder of contractile amplitudes and incresedthe contractile responses induced by EFS on gastric fundus circular muscle inhigh dose gavage and intraperitoneal injection rats (P<0.05)，while berberinelow dose gavage had no effect on the contractions (P>0.05).1,10μmol·L-1berberine increased the contractile responses concentrationdependently on gastric fundus circular muscle in normal (increase20.8%and47.7%respectively, P<0.05) and diabetic (increase54.5%and130.6%respectively, P<0.05) rats, and corrected the disorder of contractile amplitudesof gastric fundus smooth muscle in diabetic rats (improvement rate:100%). Inthe present of L-NAME only or L-NAME, α-Chymotrypsin and Suramintogether, EFS-induced contractile responses on gastric fundus circular musclecould be increased in normal and diabetic rats, and the disorder of contractileamplitudes could be corrected in diabetic rats, besides,10μmol·L-1berberineincreased further contractile responses in diabetic rats by.The contractile responses induced by EFS on gastric fundus circularmuscle were completely blocked by1μmol·L-1atropine or0.1μmol·L-1TTX(P<0.01).2.2Effects of berberine on the relaxant responses to EFS in gastric funduscircular muscle and mechanisms in vitro and in vivo administrationThe excessive enhancement of relaxant responses induced by EFS ongastric fundus smooth muscle were significantly improved in berberine highdose gavage group rats (P<0.01), however, there were no significant improvements in berberine low dose gavage and intraperitoneal injectiongroups rats (P>0.05).When the strips were pre-contracted by0.3μmol·L-1carbachol,EFS-induced relaxations were slight improved by10μmol·L-1berberine; butwhen pre-contracted by0.1μmol·L-1SP, berberine administrated in vitro hadno effects on the relaxnat responses of gastric fundus circular muscle innormal rats (P>0.05).When the strips were pre-contracted by0.3μmol·L-1carbachol,EFS-induced relaxations were inhibited significantly by100μmol·L-1L-NAME on gastric fundus circular muscle in diabetic and berberine highdose gavage group rats; the percentages of inhibition on the relaxations were93.7%and91.6%at1Hz, respectively; and there were no differences on therelaxations between two groups rats (P<0.05). The relaxations were furtherinhibited by combined L-NAME with α–chymotrypsin or suramin (P<0.05).2.3Effects of berberine on the relaxant responses to EFS in pylorus circularmuscle in vitro and in vivo administrationBerberine improved the relaxant responses on pylorus circular muscle inhigh dose gavage and intraperitoneal injection groups rats, while berberinelow dose gavage had no effect on the relaxations (P>0.05).1,10μmol·L-1berberine administrated in vitro significantly inhibitedrelaxant responses induced by EFS on pylorus circular muscle in normal anddiabetic rats (P<0.01).Conclusions1. Based on the changes of contractile and relaxant responses on gastricsmooth muscle in STZ-induced diabetic4weeks rats, the distinct sensitivity indifferent parts of stomach indicates that the damages of gastric fundus andpylorus seem to occur first on gastric functional changes in early diabetic rats.On neurogenic contractile and relaxant responses induced by EFS, thecholinergic and nitrergic nerves were damaged, and purinergic component ofthe relaxant responses to EFS was enlarged on gastric fundus strips of diabeticrats. 2. Berberine improves the functions of cholinergic and nitrergic nerve, aswell as the changes of purinergic neurotransmitter ATP in early diabetic rats,which dominated the contraction and relaxantion of gastric smooth muscle.The regulated effects of berberine on cholinergic nerve were likely to dependin part on the regulation of nitrergic nerve. Different administration routes ofberberine had different effects on gastric smooth muscle in diabetic rats; itmay involve different metabolic pathways of berberine. |
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