The Protection Of Iptakalim On Parkinson's Disease Rat Model And The Modulation Of ATP-sensitive Potassium Channels On BV-2 Cells Activation

Parkinson's disease (PD) is the most common neurodegenerative movement disorder. Clinically, most patients present with a motoric disorder and suffer from slowness of movement, rest tremor, rigidity, and disturbances in balance. The major pathological change in PD is degeneration of dopamine containing neurons of the substantia nigra pars compacta (SNpc) and the appearance of Lewy bodies. Nowadays, the main therapy has relied on dopamine supplement with levedopa (L-DOPA), with great side effects in the long run. Although many effective symptomatic therapies have been invented, there are no proven neuroprotective or neurorestorative therapies.The etiology of PD is still not fully understood, but genetic analyses, epidemiologic studies, neuropathologic investigations, and new experimental models of PD are providing important new insights into the pathogenesis of PD. PD patients have lower complex I activity, increased reactive oxygen species production, and increased radical scavenging enzyme activities, consistent with the notion that defects in complex I play a role in PD.Recently, K_ATP channels were found to have important protective functions in the hypoxia and ischemia. According to this channel, wedeveloped a new compound iptakalim, which has been demonstrated to be a novel K_ATP channel opener by pharmacological, electrophysiological, and biochemical studies. Notably, IPT is one of the KCOs passing through blood-brain-barrier, which makes it possible to investigate its effects in Parkinson's disease systematically in vivo and in vitro.Part Ⅰ Iptakalim improves rotenone related motor and neurochemical alterations in ratsAim: To investigate what kind of the role IPT plays in rotenone induced Parkinsonian rats and the underlined mechanisms.Method: In this study, PD model rats were induced by rotenone (2.5mg/kg, s.c.) for four weeks. The catalepsy test was chosen to screen the successful PD rat models. Rats were divided as (unit: mg/kg/day): Group A: vehicle + 0.2% DMSO; Group B: rotenone model rats + 0.2% DMSO; Group C: rotenone model rats + IPT 0.75; Group D: rotenone model rats + IPT 1.5; Group E: rotenone model rats + IPT 3.0; Group F: rotenone model rats + L-DOPA 10; Group G: rotenone model rats + diazoxide 3.0; Group H: rotenone model rats + 5-HD 5.0 + IPT 3.0; Group I: rotenone model rats + 5-HD 5.0 + diazoxide 3.0. The catalepsy test was used to assess the effects of L-DOPA, IPT, diazoxide on rotenone-induced Parkinsonian symptoms; HPLC-ECD was introduced for monoamine neurotransmitters assessment; RT-PCR was used to analyze iNOS mRNA level in striatum and SN; a commercially available kit was used to assay iNOS activity in striatum and SN.Result: IPT not only decreased the time of catalepsy, but increased themonoamine levels in the striatum and SN of rotenone-treated rats. IPT reduced the mRNA level and activity of iNOS in PD rats through the activation of mito-K_ATP channel. These effects of IPT could be abolished by 5-HD.Conclusion: In this study, IPT improved both Parkinsonian symptoms (e.g., dereasing the time of catalepsy) and neurochemistry alterations induced by rotenone in rats. Mito-K_ATP channel stimulators may be therapeutically used for the early PD, through its regulation of iNOS transcription and activity.Part Ⅱ the modulation of ATP-sensitive potassium Channelson BV-2 cells activationAim: To investigate the effects of IPT on rotenone induced BV-2 cells inflammatory mediator production and the possible mechanisms.Method: In this study, the murine microglial cell line BV-2 cells were introduced in this experiment. The cells were divided as: Group A: vechile; Group B: 10 nmol/L rotenone; Group C: 10 μmol/L pinacidil + 10 nmol/L rotenone; Group D: 10 μmol/L glibenclamide +10 μmol/L pinacidil +10 nmol/L rotenone; Group E: 10 μmol/L IPT +10 nmol/L rotenone; Group F: 250 μmol/L 5-HD + 10 μmol/L IPT +10 nmol/L rotenone; Group G: 100 μmol/L diazoxide +10 nmol/L rotenone; Group H: 250 μmol/L 5-HD + 100 μmol/L diazoxide +10 nmol/L rotenone.We introduced western blotting for the anal...