Development of multiple calcium channel types in cultured mouse hippocampal neurons

The development of multiple calcium channel activities was studied in mouse hippocampal neurons in culture, using the patch-clamp technique. A depolar izing pulse (40-50 ms duration) from the holding potential of - 80 mV to le vels more depolarized than - 40 mV produced a low threshold T-type current. The T-type current was observed in 52% of four days in vitro neurons. The number of neurons which expressed T-type current decreased with age of cult ure, so that the current was detected in only 18% of neurons after 16 days in vitro. The T-type current densities varied between 1.9 pA/pF and 3.29 pA /pF in the mean values during the period studied (4-16 days in vitro). A de polarizing pulse from - 80 mV to levels more depolarized than -35mV evoked a high threshold calcium channel current. The high threshold current densit y increased in the mean values from 3.9 pA/pF in four days in vitro neurons to 28 pA/pF in 16 days in vitro neurons. We have then examined the effect of nifedipine, omega-Agatoxin IVA and omega-conotoxin GVIA on the high thre shold current. Nifedipine (1-5 mu M) sensitive current density stayed in th e range of 1.9-2.1 pA/pF during 4-16 days in vitro, while omega-Agatoxin IV A (200 nM) sensitive current density increased in the mean values from 1.54 pA/pF in four days in vitro neurons to 21.5 pA/pF in 16 days in vitro neur ons. The omega-conotoxin GVIA sensitive N-type channel current was maximum at eight days in vitro (5.44 pA/pF) and it reduced progressively to reach a lmost half (2.46 pA/pF) in 16 days in vitro neurons. These results showed that diverse subtypes of calcium channels change in de nsity during the early period of culture. We suggest that the temporal expr ession of each type of channel may be linked to the development of neural a ctivities. (C) 1999 IBRO. Published by Elsevier Science Ltd.