Behavioral And In Vivo Electrophysiological Research On The Neuroprotective Effects Of Liraglutide

Alzheimer disease (AD) is a chronic, primary and irreversible neurodegenerative disorder characterized by deterioration of cognitive and mental function, including learning and memory. One of the important neuropathological features is the high density of senile plaques in the brain of AD patients. The main constituent of senile plaques is ?-amyloid protein (A?). At present, the neurotoxicity of A? has been widely reported in vivo and in vitro, including the impairment of spatial learning and memory and synaptic plasticity such as long-term potentiation (LTP). Therefore, the A? hypothesis in AD has been accepted widely. However, the possible mechanisms by which A? impairs cognitive function are complicated and unclear so far, and the effective measures in the prevention and treatment of AD are still seriously absent.It has been reported recently that AD and type2diabetes mellitus (T2DM), another degenerative disease, share several common clinical and pathological characteristics. On the one hand, T2DM is a risk factor for developing AD in the elderly. Since the impairment of insulin system in the brain is closely related to the development of neurodegenerative disorders, it might be a well promising strategy to normalize insulin signaling in the brain for the prevention and treatment of AD. One promising treatment for AD is using insulin-releasing gut hormone glucagon-like peptide-1(GLP-1), a modulator used in T2DM therapy. Interestingly, GLP-1has been reported to cross the blood brain barrier (BBB) and facilitate insulin signaling. GLP-1can reduce the levels of A? in the brain in vivo and reduce levels of amyloid precursor protein (APP) in cultured neuronal cells and possess neurotrophic properties to protect neurons against glutamate-induced apoptosis. Previous study has shown that the hippocampal LTP detrimental effect induced by A? fragments was effectively prevented by GLP-1. More importantly, GLP-1and GLP-1receptors (GLP-1R) are expressed in the brain, including the hippocampus. Another issue is the shorter half life of GLP-1, only several minutes in blood plasma, which seriously limits its application in clinical practice. Recently, Liraglutide, a novel long-lasting GLP-1analogue and greater biological activity, has been brought on the market as diabetes therapeutics. However, it is not completely understood what role the replacement of neurons plays in the brain, and whether these regenerative neurons are actually functionally integrated into the neuronal networks and the reduced A? level in the brain is related to the behavioral improvement in AD. Therefore, Liraglutide, not natural GLP-1, was used in the present study.By using the high-effective GLP-1analogue, Liraglutide, we investigated:(1) the effects of intrahippocampal injection of Liraglutide on the A?25-35-induced impairment of spatial learning and memory of rats in Morris water maze test;(2) the effects of intrahippocampal injection of Liraglutide on the A?25-35-induced impairment of in vivo L-LTP in rat hippocampal CA1region.Part ? Liraglutide Protects against A?25-35-Induced Impairment of Spatial Learning and Memory in RatsTo characterize the neuroprotective role of Liraglutide in the brain, we investigated the effects of intrahippocampal injection of Liraglutide on the A?25-35-induced impairment of spatial learning and memory of rats in a Morris water maze test. The escape latency (s), distance traveled (cm) and swimming speed (cm/s) were calculated in acquisition phase (hidden platform tests), and the percentage of the total time and the distances in the different quadrants was recorded in probe trials.The results showed that:(1) intrahippocampal injection of4nmol A?25-35impaired the spatial learning and memory of rats. In acquisition test, the latencies and distances for searching for the platform were significantly larger compared with control from the2nd to the5th training day (<(0.01). For example, the mean escape latencies and distances were17.62±1.22s and262.93±28.85cm, significantly larger (P<0.01) than the values of12.30±0.20cm and150.69±14.02cm in the control group (P<0.01) on the5th day of testing. In probe trial (6th day), the percentages of time and distance in the A?25-35group significantly decreased compared to the control group (P<0.01), with the values of24.85±0.89%and23.30±2.06%, respectively.(2) Liraglutide alone did not affect normal cognitive behavior.5nmol Liraglutide had no effects on the escape latencies and distances ((P<0.05) in hidden platform test, as well as the percentages of time and distance in the target quadrant in probe trial (P<0.05).(3) Pretreatment of Liraglutide effectively protected spatial learning and memory against A?25-35-induced impairment in a dose-dependent manner.0.05nmol Liraglutide did not affect A?25-35induced impairment of spatial learning, with similar escape latency and similar escape distance in hidden platform test as compared to the A?25-35alone group ((M0.05). However, pretreatment with0.5nmol or5nmol Liraglutide significantly reversed the spatial learning impairments induced by A?25-35(P<0.05) in most training days. For example, on the5th day of hidden platform test, the escape latencies in0.5nmol and5nmol Liraglutide plus A?25-35groups were14.50±0.36s and13.32±0.31s, respectively, showing a dose-dependent decrease. In probe trial, the percentages of total time elapsed in the target quadrant were38.01±1.67%and43.22±0.93%for0.5nmol and5nmol Liraglutide puls A?25-35group, respectively, significantly higher than A?25-35alone group (*<0.01), showing a significant dose-dependent increase (*<0.01).(4) The visible platform tests showed that all drugs did not affect the vision and the swimming speeds (>0.05).These results indicated that intrahippocampal administration of A?25-35impaired cognitive function of rats, while pretreatment with Liraglutide effectively reversed A?25-35-induced the impairment of spatial learning and memory in a dose dependent manner, suggesting that natural GLP-1in CNS may play an important positive role in maintaining normal cognitive function, and Liraglutide might be useful in the prevention and treatment of AD.Part ? Liraglutide Protects against A?25-35-Induced Impairment of Hippocampal Late-Phase Long Term Potentiation in Rat Hippocampal CA1Region in vivoIn order to clarify the mechanism by which Liraglutide protects against A?-induced behavior impairment, in vivo electrophysiological recording of L-LTP in hippocampal CA1region of rats was performed after finishing the behavioral study. Hippocampal fEPSP were recorded in the CA1region. Three sets of high frequency stimulation (HFS) with5min of interval were applied to produce a robust L-LTP.The results showed that:(1) A?25-35, not Liraglutide, significantly suppressed L-LTP. After application of4nmol A?25-35, the average fEPSP amplitudes after HFSs were133.75±6.46%,118.70±4.05%and113.50±3.19%in A?25-35group at1h,2h and3h, respectively, significantly lower than the values of162.28±4.94%,159.24±1.85%and153.41±2.79%in control group at the same time points (((<0.01). However, there was no significant change in fEPSP amplitude at1h,2h and3h after HFSs in Liraglutide alone group (5nmol) as compared to the control group.(2) Pretreatment with Liraglutide protected against A?25-35-induced impairment of L-LTP. The average fEPSP amplitudes in0.5nmol and5nmol, but not0.05nmol, Liraglutide plus A?25-35group were140.84±4.81%and152.49±4.60%at3h post-HFSs, respectively, significantly larger than the values in A?25-35alone group (P<0.05).(3) All drugs used in the experiment did not affect PPF (P>0.05)These results demonstrated that pretreatment with Liraglutide effectively protected against A?25-35-induced suppression of L-LTP in a dose-dependent manner in the rat hippocampal CA1region in vivo. The results may in part explain the cellular mechanisms of GLP-1in improving spatial cognitive function and be well consistent with the behavior experiment results above.In conclusion, the present study, using Morris water maze test and field potential recording, observed the effects of Liraglutide on the spatial learning and memory and hippocampal L-LTP in vivo, and investigated the neuroprotective effects of Liraglutide against A?25-35-induced neurotoxicity. The results indicated that Liraglutide can effectively protect against A?25-35-induced impairment of rat cognitive function and hippocampal L-LTP in vivo; the neuroprotective effect of Liraglutide may be related to the synthesis of new mRNAs and proteins of synaptic transmission. Therefore, the present experiment provides further behavioral and electrophysiological evidence for the neuroprotective effect of GLP-1analogues Liraglutide, reveals its possible cellular mechanism and its potential clinical values in the prevention and treatment of AD in the future. Objective To investigate the correlation of hippocampal synaptic plasticity with spatial learning and memory under normal and pathological condition, and provide experimental evidence for the consistence of hippocampal L-LTP and behavioral experiments. Method38SD rats were randomly divided into two groups, control and AD model. First, Morris water maze was used to test the ability of spatial learning and memory of rats. The escape latencies for rats to search for an underwater platform in5days of navigation tests and the swimming time percentage in target quadrant on the6th day after withdrawing the platform in probe trails were recorded. Then, in vivo hippocampus L-LTP of field exicitatory postsynaptic potential (fEPSP) in CA1region was recorded after delivering high frequency stimulation (HFS). Result In control group, there was a significant negative correlation between the amplitude of fEPSP and the escape latency (r=-0.8306) and a significant positive correlation between the amplitude of fEPSP and the swimming time percentage in target quadrant (r=0.7709). In AD model group, similar correlations were found, with a correlation coefficient of r=-0.7675and r=0.8049, respectively. When putting all data from the two groups together, the hippocampal L-LTP was more correlated with escape latency (r=-0.9124) and swimming time percentage (r=0.9745). Conclusion There is a very close correlation between the hippocampal L-LTP and the spatial learning and memory behavior in rats, suggesting that the hippocampal L-LTP may be involved in the electrophysiological mechanism of spatial learning and memory in rats, and the impairment of L-LTP could partly represent the deficits in cognitive function of animals such as AD.