Function And Molecular Expression Of Two Types Of Potassium Channels In Interstitial Cells Of Cajal From Diabetic Gastroparasis Rats

Interstitial cells of Cajal (ICC) are specialized cells in the gastrointestinal (GI) tract that are mesenchymal in origin and fundamental to the physiological functions of GI muscles. ICCs are proposed to have the following functions: to generate slow waves, to mediate neurotransmission between the enteric nerves and the gastrointestinal muscles and to act as mechanoreceptors. Furthermore, the abnormal changes of the number and structure of ICC have been described in a variety of gastrointestinal motility disorders.The ionic currents are involved in generation of slow waves. Voltage-dependent K+ channels (Kv channels) participate in electrical rhythmicity and smooth muscle responses by contributing to the plateau potential of slow waves and action potentials. Kv channels may also contribute to the repolarization phase of excitable events and the resting potential between slow waves. Kv channels also participate in the ICC and smooth muscle cell (SMC) responses to neural inputs because they are regulated by excitatory and inhibitory neurotransmitters. Besides,an inward rectifier K+ conductance could contribute to the negative resting potential and the membrane potential gradient across the circular muscle layer in the colon.Therefore,the dissertation aims at investigating the changes of the two types of potassium channels in ICC from normal and GDP rats by the methods of patch clamp and RT-PCR techniques. PartⅠOutward current and inward rectifier potassium current in interstitial cell of Cajal from diabetic gastroparasis ratsObjective:To explore outward current and inward rectifier potassium current in interstitial cell of Cajal from diabetic gastroparasis rats.Methods: SD rats were randomly divided into diabetic group and control group.The diabetic model of rats were duplicated with STZ once intraperitoneally. After establishing diabetic rat model for 3 months, interstitial cells of Cajal were isolated with collagenaseⅡand identified with the c-Kit antibody. Then, outward current and inward rectifier potassium current were recorded by using whole-cell patch clamp technique.Results:The I-V curve of diabetic gastroparasis group upward compared with the normal group. The peak current of outward current in diabetic gastroparasis rat raised from 641.7±59.58 pA (as control) to 1345.2±151.82 pA, at +20mV. V1/2 value of outward current in GDP rat shifted to left on abscissa about 15 mV and the differences were significant by comparison with the control, but there is no obvious change about the value of k.However,the inward rectifier potassium current of diabetic gastroparasis group decreased.PartⅡExpression of Kv1.1 and Kir2.1 mRNA in diabetic Gastroparasis ratsObjective: To explore the changes of Kv1.1 and Kir2.1 mRNA expressions at different stage of diabetic gastroparasis.Methods: Kv1.1 and Kir2.1 mRNA expressions in corpus gastricum and gastric antrum at different stage of diabetic gastroparasis(4weeks,8weeks,12weeks) were assayed by semi-quantity reverse transcription-polymerase chain reaction(RT-PCR).Results: Kir2.1/β-actin density ratio in corpus gastricum of DGP rats for 4, 8 and 12 weeks were decreased from 1.5892±0.1752 (as control) to 1.4610±0.1121, 0.8576±0.1594 and 0.5312±0.1886(P<0.05), respectively, meanwhile density ratio in gastric antrum were declined from 1.9419±0.0280 to 1.6586±0.0563, 0.6687±0.2262 and 0.5235±0.2539(P<0.05), respectively. These suggested that Kir2.1 mRNA expression in corpus gastricum and gastric antrum were time-dependently decreased in the process of DGP. Kv1.1/β-actin desity ratio at different stage of diabetic gastroparasis was almost the same.Conclusion: 1)Results showed the increase of the peak current of outward current and the decrease of the inward rectifier potassium current in diabetic gastroparasis rat and changes of these two potassium currents in ICC from diabetic gastroparesis rats probably contribute to the stability of the negative resting potential and suppress the excitability; 2)Kir2.1 mRNA expression in corpus gastricum and gastric antrum were time- dependently decreased in the process of DGP. It indicated that the number of Kir2.1 in corpus gastricum and gastric antrum decreased gradually in the process of DGP.These findings probably indicated that changes of function and molecular expression of two types of potassium channels in interstitial cells of Cajal are one of the mechanisms of gastroparesis in diabetic rats.