VOLTAGE-CLAMP ANALYSIS OF MOSSY FIBER SYNAPTIC INPUT TO HIPPOCAMPAL-NEURONS
VOLTAGE-CLAMP ANALYSIS OF MOSSY FIBER SYNAPTIC INPUT TO HIPPOCAMPAL-NEURONS
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DOI：
10.1152/jn.1983.50.2.487
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发表时间：
1983-01-01
影响因子：
2.5
通讯作者：
JOHNSTON, D
中科院分区：
文献类型：
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作者：
BROWN, TH;JOHNSTON, D
用离体脑片法研究了海马锥体神经元诱发的突触反应。使用电流钳和电压钳技术，突触反应的幅度和时间过程被测量为突触后细胞膜电位的函数。这些结果首次描述了哺乳动物皮层神经元中负责产生突触信号的突触后传导机制。细胞内记录用低电阻（10 - 30 M Ω.）玻璃微量移液管，其通常填充有2 M Cs2SO4或4 M乙酸钾。在直接视觉控制下，将记录微量移液器尖端定位到CA3区的锥体细胞层（锥体细胞体层）中，并将双极刺激电极放置到齿状回的颗粒层（颗粒细胞体层）中。低强度电流被用来刺激颗粒细胞，其苔藓状纤维轴突被称为使单突触传递突触到锥体神经元的近端顶端树突上。3 kHz分时单电极钳（SEC）用于电流钳和电压钳实验。在许多实验中，Cs+被注入突触后锥体神经元，以增加它们的输入电阻，减少向外整流。建立了严格的标准，接受电压钳结果，标准是在298个研究细胞中只有12个得到满足。在电流钳条件下，当突触后锥体细胞的膜电位为-60~-70 mV或更负时，突触后电位（PSP）表现为简单的单相去极化。当锥体细胞通过外向电流去极化时，PSP变为双相，由去极化早期相和超极化晚期相组成。早期相被认为代表了对颗粒细胞苔藓纤维输入的单突触反应，而晚期相被认为是由反复或前馈突触抑制引起的。当锥体细胞进一步去极化时，PSP的超极化相振幅显著增加，去极化相振幅减小。当这样的细胞去极化时，-30 mV时，整个PSP波形通常表现为超极化。当通过向浴中加入印防己毒素（5 - 10 μ M）或青霉素（3.3mM）来阻断晚期时，剩余PSP的反转电位以正方向移动到平均值（±）。SE）值为-0.3. ±-。2.9 MV.由颗粒细胞刺激产生的突触信号产生来自电导增加机制，并且电流显示出明确的反转电位，与突触传递的化学模式一致。电导波形和电流反转电位的这种表征对于理解突触信号的起源及其在整个树突状分支中的传播是重要的。分离和量化负责兴奋性和抑制性电流的电导的能力，将允许测试几种可能的机制，这些机制可能是海马中使用依赖性突触可塑性的有趣形式的基础。
Evoked synapatic responses were studied in hippocampal pyramidal neurons, using the in vitro slice preparation. Using both current- and voltage-clamp techniques, the amplitude and time course of the synaptic responses were measured as a function of the membrane potential of the postsynaptic cell. The results provide the 1st description of the postsynaptic conductance mechanisms responsible for the generation of synaptic signals in a mammalian cortical neuron. Intracellular recordings were made with low-resistance (10-30 M.OMEGA.) glass micropipettes, which were usually filled with 2 M Cs2SO4 or 4 M potassium acetate. Under direct visual control, the recording micropipette tips were positioned into the stratum pyramidale (the pyramidal cell body layer) of the CA3 region, and bipolar stimulating electrodes were placed into the stratum granulosum (granule cell body layer) of the dentate gyrus. Low-intensity current was used to stimulate the granule cells, whose mossy fiber axons are known to make monosynaptic en passant synapses onto the proximal apical dendrites of the pyramidal neurons. A 3-kHz time-share single-electrode clamp (SEC) was used for both current- and voltage-clamp experiments. In many of the experiments, Cs+ were injected into the postsynaptic pyramidal neurons to increase their input resistance and reduce outward rectification. Strict criteria were established for accepting voltage-clamp results, criteria that were satisfied in only 12 of the 298 cells studied. Under current-clamp conditions, when the membrane potential of the postsynaptic pyramidal cell was -60 to -70 mV or more negative, the postsynaptic potential (PSP) typically appeared to consist of a simple monophasic depolarization. When the pyramidal cell was depolarized by passing outward current, the PSP became biphasic, consisting of a depolarizing early phase followed by a hyperpolarizing late phase. The early phase is thought to represent the monosynaptic response to the granule cell mossy fiber input, while the late phase is believed to arise from recurrent or feedforward synaptic inhibition. When the pyramidal cell was further depolarized, the amplitude of the hyperpolarizing phase of the PSP greatly increased and the amplitude of the depolarizing phase decreased. When such cells were depolarized more positive that .apprx. -30 mV, the entire PSP waveform typically appeared to become hyperpolarizing. When the late phase was blocked pharmacologically by adding picrotoxin (5-10 .mu.M) or penicillin (3.3 mM) to the bath, the reversal potential of the remaining PSP shifted in a positive direction to a mean (.+-. SE) value of -0.3 .+-. 2.9 mV. Synaptic signal generation produced by granule cell stimulation results from a conductance increase mechanism and the currents display a clear reversal potential, consistent with a chemical mode of synaptic transmission. This characterization of the conductance waveforms and current reversal potentials is important for understanding the genesis of synaptic signals and their propagation throughout the dendritic arborization. The ability to separate and quantify the conductances responsible for the excitatory and inhibitory currents will permit tests of several of the possible mechanisms that may underlie the interesting forms of use-dependent synaptic plasticity in the hippocampus.