The influence of acetoacetate and butyrate on calcium influx and ATP concentrations in HT-29 cells.
The effects of acetoacetate and butyrate on Ca(2+)-influx in HT-29 cells were unknown. Extracellular signals can be transferred to the intracellular environment of the cell via changes in the Ca(2+)-concentration. Extracellular Ca2+ may enter the cell via Ca(2+)-channels in the plasma membrane. Physiological processes occurring within the cell are dependent on Ca(2+)-concentration, including enzyme activity. Intracellular Ca(2+)-concentrations were measured using Fura-2/AM, a fluorescent intracellular Ca(2+)-probe. Ca(2+)-concentrations were measured immediately on application of the inducers to the cells, as well as after a 9 day incubation period. The effect of these inducers on the L-type voltage-operated Ca(2+)-channels were determined using the whole-cell patch-clamp technique. To validate these results for the intestinal epithelial model, membrane current studies were performed on HT-29 cells grown on a polycarbonate membrane. ATP concentrations were measured, and the theoretical effect of the inducers on PDE 4 activity was determined. It was found that both acetoacetate and butyrate blocked Ca(2+)-influx through the L-type voltage-operated Ca(2+)-channels, resulting in the initial low Ca(2+)-concentration (p < 0.05). The blockage effect is short-lived as after a 9 day incubation period in the presence of the inducers, Ca(2+)-concentrations were higher than that of the HT-29 control sample (p < 0.05). ATP concentrations of the cells were decreased in the presence of the inducers (p < 0.05), whilst it was suggested that no interaction between the catalytic site of PDE 4 and the inducers existed.
Jamie
H
H
Cyclic Peptide Research Unit, Potchefstroom University for C.H.E., South Africa.
Dyason
K
K
Milne
P J
PJ
Grant
G
G
Graz
C J
CJ
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000378:metabolism; D000090:Acetoacetates / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D002087:Butyrates / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D019295:Computational Biology; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; D004790:Enzyme Induction / Q000187:drug effects; D019073:HT29 Cells; D006801:Humans; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D008958:Models, Molecular; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D018408:Patch-Clamp Techniques
D000090:Acetoacetates; D002087:Butyrates; D020746:Calcium Channels, L-Type; C016635:acetoacetic acid; D000255:Adenosine Triphosphate; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; D002118:Calcium