Patch-clamp studies in the CNS illustrate a simple new method for obtaining viable neurons in rat brain slices: glycerol replacement of NaCl protects CNS neurons.
Viable neurons in brain slices are crucial for electrophysiological studies. The present study describes a new method for obtaining viable cells in several regions of the central nervous system including the ventral tegmental area, the hypothalamus, the periaqueductal grey matter and the spinal cord. The essence of the method was to use a modified artificial cerebrospinal fluid (ACSF) in which all NaCl was replaced initially by equi-osmotic glycerol. This modified glycerol-based ACSF was used during slice preparation. The underlying principle for the modification is to prevent the possible acute neurotoxic effects of passive chloride entry, subsequent cell swelling and lysis. This method significantly increased the live/dead ratio in morphology compared to the normal ACSF or sucrose-base ACSF, in which NaCl was replaced by sucrose. An examination of some electrophysiological and pharmacological properties of the neurons in these preparations, by means of current-clamp and voltage-clamp recordings, revealed similar properties of those neurons obtained with the traditional ACSF method. Due to the increase in the number of viable neurons, the new ACSF increases the productivity of experiments. Based on our data, we propose that this glycerol-based solution may protect CNS neurons.
Ye
Jiang Hong
JH
Department of Anesthesiology, New Jersey Medical School (UMDNJ), 185 South Orange Avenue, Newark, NJ 07103-2714, USA. ye@umdnj.edu
Zhang
Jingli
J
Xiao
Cheng
C
Kong
Jian-Qiang
JQ
D000818:Animals; D000891:Anti-Infective Agents, Local / Q000494:pharmacology; D001921:Brain / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D016923:Cell Death / Q000187:drug effects; D002469:Cell Separation; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002492:Central Nervous System Depressants / Q000494:pharmacology; D002555:Cerebrospinal Fluid / Q000737:chemistry; D004566:Electrodes; D004594:Electrophysiology; D000431:Ethanol / Q000494:pharmacology; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects; D005990:Glycerol / Q000494:pharmacology*; D006096:Gramicidin / Q000494:pharmacology; D066298:In Vitro Techniques; D009046:Motor Neurons / Q000187:drug effects; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques; D051381:Rats; D017207:Rats, Sprague-Dawley; D012965:Sodium Chloride / Q000494:pharmacology*; D013116:Spinal Cord / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*
D000891:Anti-Infective Agents, Local; D002492:Central Nervous System Depressants; D006096:Gramicidin; D000431:Ethanol; D012965:Sodium Chloride; D005990:Glycerol