Electrical activity recording method for tiny single synaptic neurons

A single-synapse and neuron technology, applied in the scientific field, can solve problems such as the inability to distinguish electrical signals, and achieve the effect of simple and easy-to-use real-time detection methods and techniques

Inactive Publication Date: 2017-04-19
INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But one of the big drawbacks is the inability to distingu...

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  • Electrical activity recording method for tiny single synaptic neurons
  • Electrical activity recording method for tiny single synaptic neurons
  • Electrical activity recording method for tiny single synaptic neurons

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Experimental program
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Embodiment 1

[0046] Embodiment 1: the preparation of brain slice:

[0047] After the surgical instruments are prepared, decapitate the mice of P2-3 (2 days to 3 days after birth), and remove the brainstem with surgical scissors, ophthalmic scissors, scalpel and tweezers, and dip into the Slicing solution from time to time during this process (125 sodium chloride, 25 sodium bicarbonate, 3 inositol, 2 sodium pyruvate, 2.5 potassium chloride, 1.25 sodium dihydrogen phosphate, 0.4 ascorbic acid, 25 glucose, 0.1 calcium chloride, 3 magnesium chloride (mM, sigma), pH 7.4, with 95% oxygen and 5% carbon dioxide). Then the tissue was glued to the slicing chamber, and the brain slices with a thickness of 200 μM were cut out with a microtome. According to the age of the mouse, we can cut 1-3 brain slices containing Medial Nucleus of the Trapezoid Body (MNTB), and the process of cutting the brain slices is also carried out in this slicing solution. Keep on ice. Low Ca levels during brain slice prep...

Embodiment 2

[0048] Example 2: Whole-cell clamping

[0049] After the brain slices were incubated, quickly put a piece of brain slice on the glass slide, and cover the brain slice with a platinum U-shaped board wrapped with uniform and equidistant nylon threads (Johnson&Johnson REACH), so that the brain slice was in the There will be no movement on the slide. During the whole experiment, the brain slice was perfused with extracellular fluid (Bath solution) at a rate of 1 ml / min. Also make sure that the space above the slide does not allow extracellular fluid to overflow. Then observe with a microscope, first find the calyx of Held synapse and post-synaptic cell body with a 5× low-power lens, and then observe with a 60× diving lens. After finding the cell body with tiny monosynaptic presynaptic cells, this was clamped whole-cell with a glass microelectrode on one side, and the subsequent recording mode was performed by the MultiCalmp 700B amplifier.

Embodiment 3

[0050] Example 3: Loose-patch clamping

[0051] After the whole-cell pattern is formed, use the glass microelectrode on the other side to perform loose-patch on the observed tiny monosynaptic presynaptic cells, and the whole process should be very gentle (such as figure 2 a, b shown). Simultaneous recordings of loose patch-clamp channels and whole-cell channels are then possible. Such as figure 2 As shown in c, the signals in the upper figure are from the loose patch-clamp channel and the whole-cell channel respectively, which contain the signal of micro-post-synaptic current (mEPSC); Positive-going electrical signals in patch-clamp recording mode were only highly time- and amplitude-correlated with certain mEPSCs in whole-cell recordings (e.g. figure 2 d), which shows that the recorded mEPSC signal originates from this tiny monosynaptic junction.

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Abstract

The invention discloses an electrical activity recording method for tiny single synaptic neurons and belongs to the field of neurobiology research. Based on the conventional patch clamp technique, electrical signals input through all tiny presynaptic neurons can only be recorded through postsynaptic neurons, while electrical signals input by each tiny presynaptic neuron cannot be recognized. In this way, the invention provides the above method. According to the technical scheme of the invention, the loose-patch clamp technique and the whole-cell patch clamp technique are integrated, and the electrical activity of each tiny single synapsis can be recorded simply on a brain slice in the physiological state. Therefore, based on the method, the dynamic characteristics of the vesicle spontaneous release and the vesicle induced release of the single synapsis can be detected. The method provides a direct, quantitative, real-time and high-time-resolution detection method for the dynamic characteristics of the vesicle release of the single synapsis and the synaptic plasticity.

Description

technical field [0001] The invention belongs to the field of scientific and technical methods, and specifically relates to a method for studying the kinetic characteristics of single-synaptic vesicle release under physiological conditions and providing a direct, quantitative, real-time and high-time-resolution method for single-synaptic plasticity. Detection method. In particular, it relates to an electrophysiological detection method for the research in the field of nerve synapse transmission and the research of synapse-related neurological diseases. Background technique [0002] As early as the 1950s, when Paul Fatt and Bernard Katz were studying the neuromuscular junction (NMJ) of frogs, they discovered that even if neurons were not activated to generate action potentials, they could also record spontaneous properties from muscle cells. The tiny endplate potential (spontaneous miniature endplate potential, mEPP). This spontaneous nature of tiny potentials has since been...

Claims

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Application Information

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IPC IPC(8): G01N33/483
CPCG01N33/4836
Inventor 戴金叶孙坚原陈培华张树利
Owner INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES
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