Effects Of Nicotine On Tactile Stimulation-evoked Synaptic Transmission And Plasticity At Mouse Cerebellar Granule Cell Layer

[Purpose]Nicotinic acetylcholine receptors(nAChRs)are cationic channels that mediate fast excitatory transmission in the central nervous system.Several nAChR subunits have been detected within cerebellar granule cell layer(GCL),and activation of nAChRs may have a significant influence on neuronal synaptic transmission of the cerebellum.The mammalian cerebellum is a critical organ that controlling motor coordination,planning and fine regulation of voluntary movement,it is also very important for cognitive function,while nicotine modulates neuroplasticity and improves working memory performance,episodicmemory performance and motor functions by modulating long-term depression and long-term potentiation.The aim of present study was to better understand the roles of nAChRs during the sensory stimulation-evoked synaptic transmission in the cerebellar GCL,and study is to investigate the effect of nicotine on the facial stimulation-evoked long-term synaptic plasticity in cerebellar granule cell layer in vivo in mice.[Methods]A total of 42(6-8-week-old)ICR mice were used in this study.After the mice were anesthetized with urethane(1.3 g/kg body weight i.p.),the craniotomy was drilled to expose the cerebellar surface corresponding to Crus ?.The cerebellar surface was constantly superfused with oxygenated artificial cerebrospinal fluid(ACSF)with a peristaltic pump at 0.5 ml/min.Rectal temperature was monitored and maintained at 37.0 ± 0.2 ? using body temperature equipment.Local field potential recordings from GCL were performed with an Axopatch-200B amplifier(Molecular Devices,Foster City,USA)under current-clamp recording mode(I=0),at depths 350-450 ?m under pia mater membrane,which were confirmed from the display of the micro-manipulator(MP-285).The signals were filtered at 5 kHz,and acquired through a digidata 1440 series analog-to-digital interface on a personal computer using Clampex 10.3 software.Facial stimulation was performed by air-puff of the ipsilateral whisker pad through a 12-gauge stainless steel tube connected with a pressurized injection system.Gabazine(20 ?M)was routinely included in external recording solutions to block Golgi cell-mediated GABAergic inhibition in the GCL.For the induction of mossy fiber-granule cell synaptic plasticity,air-puff stimulus at 20 Hz(240 pulses)were delivered 10 min after the recording became stable.The amplitude of N1 before and after 20 Hz facial stimulation was normalized by the mean value of baseline.Values are expressed as the mean ± S.E.M.One-way ANOVA(posthoc multiple comparison)and Tow-way ANOVA(SPSS software)was used to determine the level of statistical significance among groups of data.P-values below 0.05 were considered statistically significant.[Results](1)Facial stimulation induced a pair of negative components,N1 and N2 in the GCL of cerebellar cortex folium Crus ?,which identified as stimulus onset response(Ron)and stimulus offset response(Roff).Cerebellar surface perfusion of nicotine for 5 min did not significantly change the latency of the response,but induced time-dependent increases in the amplitude and area under curve(AUC)of N1.The nicotine-induced increase in the AUC of N1 was concentration-dependent.The lowest concentration of nicotine for increasing the AUC was 1 ?M.The 50%effective concentration(EC50)of nicotine was 32.6 ?M.(2)Perfusion of a specific a4?2 subunits of nAChRs,DH?E(1 ?M)neither significantly change the amplitude of N1,nor block the nicotine-induced increases in amplitude and AUC of N1.Perfusion of a7 subunits of nAChRs,MLA(1 ?M)did not significantly change the amplitude of the facial stimulation evoked N1,and failed to completely block the nicotine-induced increases in amplitude and AUC of the facial stimulation-evoked N1.However,perfusion of a mixture of DH?E and MLA did not significantly change the amplitude of the facial stimulation evoked N1 but completely prevented the nicotine-induced increases in amplitude and AUC of the facial stimulation-evoked N1.(3)The repetitive facial stimulation at 20 Hz(240 pulses)produced a LTP of field potential response in mouse cerebellar granule cell layer,which expressed long-term increases in amplitude and area under curve(AUC)of N1.Delivering the facial stimulation accompanied with application of nicotine(1 ?M)induced a stronger LTP of field potential response than control group,which expressed a strong time-dependent increase in amplitude of N1 than control group.(4)Blockade ?7 subunits of nAChRs failed to prevent the nicotine-induced enhancement of the LTP of field potential response in mouse cerebellar granular cell layer.Blockade ?4?2 subunits of nAChRs abolished the nicotine-induced facilitation of LTPof field potential response in mouse cerebellar granular cell layer.But GABAA receptor blocker failed to prevent the nicotine produced facilitation of the LTP of field potential response in mouse cerebellar granular cell layer.[Conclusions](1)Nicotine modulates the activity of both ?7 and ?4?2 nAChR subunits,resulting in an enhancement of facial stimulation-evoked responses in the mouse cerebellar GCL.(2)Nicotine facilitates the facial stimulation-produced LTP via ?4?2 subunits of nAChRs,but not ?7 subunits of nAChRs and GAB AA receptors.(3)Our results suggest that nicotine modulates sensory information processing in the cerebellar GCL through different subunits nAChRs.