Enhancement of asynchronous and train-evoked exocytosis in bovine adrenal chromaffin cells infected with a replication deficient adenovirus.
Bovine adrenal chromaffin cells share many characteristics with neurons and are often used as a simple model system to study ion channels and neurotransmitter release. We infected bovine adrenal chromaffin cells with a replication deficient adenovirus that induces expression of the common reporters beta-galactosidase and Green Fluorescent Protein via a bicistronic sequence. In perforated-patch recordings performed 48-h postinfection, peak calcium currents were reduced 32%, primarily due to loss of omega-conotoxin-GVIA-sensitive current. In contrast, sodium currents were increased 17%. Exocytosis, detected as an increase in membrane capacitance immediately after a single step depolarization, was reduced in proportion to the decrease in calcium influx. However, capacitance continued to increase for seconds after the depolarization. The amplitude of this poststimulus drift, or asynchronous exocytosis, was approximately three times that which occurred in a small fraction of control cells. Exocytosis evoked by repetitive stimulation with a train of brief depolarizations was increased 50%. Intracellular calcium levels measured during and after stimulation were lower, not higher, in adenovirus-infected cells. Electroporated cells showed reduced calcium currents but no enhancement of exocytosis. Cells infected with UV-irradiated virus showed reduced calcium currents and enhancement of exocytosis, but the changes were smaller than those caused by intact virus. Our results are consistent with the idea that adenovirus capsid and adenoviral DNA contribute to a Ca2+ influx- and [Ca2+]i-independent enhancement of exocytosis in bovine chromaffin cells.
Thiagarajan
Ramachandran
R
Dept. of Physiology, Emory University School of Medicine, Atlanta, Georgia, USA.
Wilhelm
Jennifer
J
Tewolde
Teclemichael
T
Li
Yingjie
Y
Rich
Mark M
MM
Engisch
Kathrin L
KL
D000200:Action Potentials / Q000502:physiology*; D000256:Adenoviridae / Q000502:physiology*; D000311:Adrenal Glands / Q000502:physiology / Q000821:virology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000502:physiology* / Q000821:virology*; D005089:Exocytosis / Q000502:physiology*; D008564:Membrane Potentials / Q000502:physiology*; D014162:Transfection / Q000379:methods; D038301:Virus Inactivation; D014779:Virus Replication / Q000235:genetics
D002118:Calcium