Marco Pirazzini
  • Department of Biomedical Sciences, University of Padova, Italy, Italy,
研究方向
  • Neuroscience
Preparation of Cerebellum Granule Neurons from Mouse or Rat Pups and Evaluation of Clostridial Neurotoxin Activity and Their Inhibitors by Western Blot and Immunohistochemistry
用老鼠或鼠仔制备小脑颗粒神经元并通过免疫印迹和免疫组织化学法评估梭菌神经毒素活性及其抑制剂
作者:Domenico Azarnia Tehran and Marco Pirazzini日期:07/05/2018,浏览量:9978,Q&A: 0
Cerebellar Granule Neurons (CGN) from post-natal rodents have been widely used as a model to study neuronal development, physiology and pathology. CGN cultured in vitro maintain the same features displayed in vivo by mature cerebellar granule cells, including the development of a dense neuritic network, neuronal activity, neurotransmitter release and the expression of neuronal protein markers. Moreover, CGN represent a convenient model for the study of Clostridial Neurotoxins (CNT), most notably known as Tetanus and Botulinum neurotoxins, as they abundantly express both CNT receptors and intraneuronal substrates, i.e., Soluble N-ethylmaleimide-sensitive factor activating protein receptors (SNARE proteins). Here, we describe a protocol for obtaining a highly pure culture of CGN from postnatal rats/mice and an easy procedure for their intoxication with CNT. We also illustrate handy methods to evaluate CNT activity and their inhibition.
Electrophysiological Recordings of Evoked End-Plate Potential on Murine Neuro-muscular Synapse Preparations
电生理记录小鼠神经肌肉突触制备物中诱发的终板电位
作者:Giulia Zanetti, Samuele Negro, Aram Megighian and Marco Pirazzini日期:04/20/2018,浏览量:11231,Q&A: 0
Neuromuscular junction (NMJ) is the specialized chemical synapse that mediates the transmission of the electrical impulse running along motor neuron axons to skeletal muscle fibers. NMJ is the best characterized chemical synapse and its study along many years of research has provided most of the general knowledge of synapse development, structure and functionality.

Electrophysiology is the most accurate experimental procedure to study NMJ physiology and it largely contributed to the elucidation of synaptic transmission basic principles. Many electrophysiological techniques have been developed to study NMJ physiology and physiopathology. In this paper, we describe an ex vivo tissue preparation for electrophysiology that can be applied to investigate nerve-muscle transmission functionality in mice. It is routinely used in our laboratory to study presynaptic neurotoxins, antitoxins, and to monitor NMJ degeneration and regeneration. This is a broadly applicable technique which can also be adopted to investigate alterations of NMJ activity in mouse models of neuromuscular diseases, including peripheral neuropathies, motor neuron disorders and myasthenic syndromes.
Mouse Phrenic Nerve Hemidiaphragm Assay (MPN)
小鼠中膈神经偏侧膈分析(MPN)
作者:Giulia Zanetti, Samuele Negro, Marco Pirazzini and Paola Caccin日期:03/05/2018,浏览量:9004,Q&A: 0
The neuromuscular junction (NMJ) is the specialized synapse by which peripheral motor neurons innervate muscle fibers and control skeletal muscle contraction. The NMJ is the target of several xenobiotics, including chemicals, plant, animal and bacterial toxins, as well as of autoantibodies raised against NMJ antigens. Depending on their biochemical nature, the site they target (either the nerve or the muscle) and their mechanism of action, substances affecting NMJ produce very specific alterations of neuromuscular functionality.

Here we provide a detailed protocol to isolate the diaphragmatic muscle from mice and to set up two autonomously innervated hemidiaphragms. This preparation can be used to study bioactive substances like toxins, venoms and neuroactive molecules of various origin, or to measure the force of skeletal muscle contraction.

The ‘mouse phrenic nerve hemidiaphragm assay’ (MPN) is an established model of ex vivo NMJ and recapitulates the complexity of neuromuscular transmission in a system easy to control and to manipulate, thus representing a valuable tool to study both NMJ physiology and the mechanism of action of toxins and other molecules acting at this synapse.