Study On The Effect Of Subthreshold Neuronal Activity On Synaptic Plasticity In Rat Visual Cortex

ObjectivesThe core basis of the visual development is a process of synaptic modification dependent on the visual experience in the visual system, especially the visual cortex. The traditional theory on the synaptic plasticity which is called Hebb’s rule emphasised the frequency of the presynaptic neural activities. Although the current development of this theory, the spike-timing dependent plasticity (STDP) emphasises the importance of the timing of pre-and postsynaptic neural activities, the postsynaptic activities must be suprathreshold. Back-propagating action potentials or dendrite spikes are regarded as the pre-requisite condition of the STDP rule. However, the subthreshold activity is a more common form of neuronal activity. Here we investigated the effect of the subthreshold neuronal activities on the synaptic plasticity in rat visual cortex during the critical period of visual development and to obtain further understanding on the synaptic plasticity of visual cortex.Methods1. Wistar rats of postnatal19-14days were used for this study. By combining infrared differential interference contrast (ID-DIC) technique and CCD-camera system, whole-cell recording were done using visualized patch clamp technique.1) The patch electrode was put in the layer IV of visual cortex. Bipolar stimulus electrode was put in the white matter corresponding to the recorded position. After modulating the stimulus intensity of the bipolar electrode, a mild subthreshold excitatory postsynaptic current (EPSC) was obtained from the postsynaptic neuron in layer IV of visual cortex.2) Under current clamp mode, a step current injection was done on the postsynaptic neuron and the threshold stimulus intensity was determined. The stimulus intensity with one step lower than the threshold intensity was used for the postsynaptic stimulus for training condition.3) The pre-and postsynaptic paired stimulus was used for training condition which was controlled by a program and with different pre-and postsynaptic timing intervals (At) and fixed frequency of1Hz and repeated for100times. 2. The following training stimuli were used, respectively. The EPSC before and after training were determined for observation of the synaptic transmission efficay changes.1) For study on the synaptic plasticity direction and timing window, a paired protocol with postsynaptic subthrehold stimulus of one step lower than the threshold intensity with timing intervals of-230ms-+230ms, postsynaptic stimulus duration of30ms was used.2) For study on the synaptic plasticity of the subthreshold stimulus level, a paired protocol with postsynaptic subthrehold stimulus of half of the threshold intensity with fixed At was used.3) For study on the synaptic plasticity of postsynaptic stimulus duration, a paired protocol with postsynaptic subthrehold stimulus of one step lower than the threshold intensity with fixed At, and postsynaptic stimulus duration of5ms,10ms,15ms,20ms,25ms,30ms, respectively was used.4) For study on the synaptic plasticity of postsynaptic burst stimulus, a paired protocol with postsynaptic subthrehold stimulus with duration of single5ms and burst form of double5ms with5ms interval was used.5) For study on the synaptic plasticity of inhibited membrane repolarization, paired protocols of pre-post mode (△t>0) and post-pre mode (△t<0) were used. Potassium channel blocking agents CsCl or TEA and combined with voltage-gated calcium channel agent NiCl2were used to observe their effects.3. pClamplO and Matlab2008were used for analysis and data processing. Origin8.0was used for statistical analysis. The directions, amplitudes, timing window of synaptic plasticity induced by subthreshold depolarization paired stimulus training were observed, respectively. The effect of depolarization level, duration and burst form on the synaptic plasticity were observed. The pharmacological effects of TEA and CsCl and NiCl2on synaptic plasticity were observed.Results1. Study on the directions and timing window of synaptic plasticity induced by subthreshold neuronal activities.1) Under△t=+10ms, LTP were induced significantly, the normalized EPSC amplitudes before training and after training at60min (t=2.94, P<0.01) and30min (t=2.79, P<0.05) were significantly different.2) Under At=+70ms, LTD were induced significantly, the normalized EPSC amplitudes before training and after training at60min (t=2.79, P<0.05) and30min (t=2.62, P<0.05) were significantly different.3) Under At=-70ms, LTD were induced significantly, the normalized EPSC amplitudes before training and after training at60min (t=3.62, P<0.015) and30min (t=2.70, P<0.05) were significantly different.4) The timing window in which LTP or LTD can be induced is At=-140ms-+140ms, which can be divided into three sub windows, zone I of At<0inducing LTD, zone Ⅱ of At0～+50m inducing LTP, zone Ⅲ of△t>+70ms inducing LTD.5) There is no dependence of synaptic plasticity on initial synaptic strength in zone Ⅰ and Ⅲ. hi zone Ⅱ, with the increasing of synaptic strength, the changing amplitude of synaptic efficacy was decreased.6) In zone I, under medium and high excitabilities, the synaptic efficacy change increased with decreased excitability, that is to say the decreased excitability induced greater LTD. While under low excitability, lower excitability induced less LTD and even LTP. In zone Ⅱ, under high excitability, the synaptic efficacy inclined to LTD with decreased excitability. While under medium and low excitability, lower excitability induced more LTP. In zone Ⅲ, the synaptic efficacy inclined to LTD with decreased excitability.2. Study on the effect of subthreshold depolarization level, duration and burst form on the synaptic plasticity.1)Under At=+40ms and duration=30ms, low level subthrehold deporalization significantly reversed the LTP effect (t=2.43, P<0.05). Under At=-40ms and duration=30ms, low level subthrehold deporalization inclined to induce lower LTD with no significantly different compared with control (t=0.55, P>0.05).2)Under At=+40ms,5m duration subthrehold deporalization significantly reversed the LTP effect under30ms duration (t=2.25, P<0.05).3) Under different durations of At=+5ms-30ms, with the shortened duration, the direction of the synaptic plasticity was reversed progressively. LTP was induced under duration exceeding20ms and LTD was induced under duration shorter than20ms. Under At=-40ms, low level subthreshold depolarization inclined to induce less LTD with no significantly difference (t=0.64, P>0.05).4) Under At=+40ms5ms duration depolarization burst significantly reversed the LTD induced by single5ms depolarization into LTP (t=2.46, P<0.05). There were also differences among conditions of5ms duration depolarization burst and single5ms depolarization and single15ms depolarization (ANOVA, F=5.2, P<0.01;t(10ms:5ms burst)=2.77, P<0.05;t(15ms:5ms burst)=2.79, P<0.05).3. Study on the effect of inhibition of cellular membrane repolarization on the subthreshold neuronal activity induced visual synaptic plasticity.1) Under△t=+10ms, CsCl and TEA both caused significantly LTD compared with control(ANOVA, F=9.74, P<0.001;t(control:CsCl)=4.07, P<0.01;t(control:TEA)=3.32, P<0.01).2) Under△t=-10ms, CsCl caused significantly LTD dependent on depolarization duration. There was significantly different under prolonged depolarization (ANOVA, F=5.48, P<0.05;t(control:CsCl5ms)=2.08, P=0.06;t(control:CsCl30ms)=3.22, P<0.01).3) Voltage-gated channel blocking agent NiCl2significantly reversed the effect of TEA and the synaptic efficacy recovered to the control level (ANOVA, F=9.92, P<0.01;t(control:TEA)=2.16, P<0.01;t(TEA:TEA+CsCl)=4.42, P<0.01).Conclusions1. Subthreshold neuronal activities changed the synaptic transmission efficacy in a timing window of△t=-140ms～+140ms. The profile of this kind of bidirectional synaptic plasticity is asymmetical similarly to that of STDP. An additional LTD window exists in rat visual cortex in subthreshold activitiy induced timing-dependent synaptic plasticity.2. The initial synaptic strength and the neuronal activity can modulate the the bidirectional synaptic plasticity induced by subthreshold neuronal activity. Their modulating effects are different dependent on timing window zone.3. The depolarization level, duration, and burst form can modulate the bidirectional synaptic plasticity induced by subthreshold neuronal activity. Low level subthreshold depolarization and short duration induce more LTD. Burst form induced more LTP.4. The activation of voltage-gated calcium channel play a key role in determination of the direction of synaptic plasticity induced by subthreshold neuronal activity. 5. The postsynaptic action potential is not the prerequisite condtion for the timing-dependent plasticity in visual cortex.