Acute isolation of caudate nucleus neurons with patch-clamp technique in neonate rats / 中国组织工程研究

ABSTRACT

BACKGROUND: Single scattered neurons are needed in most studies on ion channels. However, artificial primary cultured neurons, for the influence from the environment, have a lot of changes in their characteristics. Comparatively, acutely isolated neurons can keep intact physiological characteristics. OBJECTIVE: To establish an ideal method for acute isolation of caudate nucleus neurons and to study their ion channels and signal transduction mechanism with patch clamp technique for the benefit of deeply understanding functions of caudate nucleus. DESIGN, TIME AND SETTING: An observational experiment on cells was performed at the Central Laboratory, Medicine Science College of China Three Gorges University from February to December in 2008. MATERIALS: Twelve 7- to 10-day-old neonatal Wistar rats, of either gender, were selected for this study. METHODS: Scattered single caudate nucleus neuron was obtained from 7- to 10-day-old neonatal Wistar rats with a combination of enzymatic and mechanical separation method. Voltage-dependent calcium currents were determined with whole-cell patch clamp technique MAIN OUTCOME MEASURES: Observation of morphology and electrophysiological characteristics of acutely isolated caudate nucleus neurons with whole-cell patch clamp. RESULTS: The caudate nucleus neuron acutely isolated with protease digestion and mechanical isolation method was found to have regular configuration, intact cell membrane, long neuraxons, excellent morphology and physiological nature. Their L-calcium ion channel current observation showed electrical parameters within normal physiological range, and activity of voltage-dependent calcium ion channel was well preserved. CONCLUSION: As neurons isolated in this way have normal morphology and long neuraxons, and activities of primary iron channels are well preserved, we conclude that an acute isolation method of caudate nucleus neurons from neonate rats with patch clamp technique is successfully established.