Molecular and functional expression of high conductance Ca 2+ activated K+ channels in the eel intestinal epithelium.
Several types of K(+) channels have been identified in epithelial cells. Among them high conductance Ca(2+)-activated K(+) channels (BK channels) are of relevant importance for their involvement in regulatory volume decrease (RVD) response following hypotonic stress. The aim of the present work was to investigate the functional and molecular expression of BK in the eel intestine, which is a useful experimental model for cell volume regulation research. In the present paper using rat BK channel-specific primer, a RT-PCR signal of 696 pb cDNA was detected in eel intestine, whole nucleotide sequence showed high similarity (83%) to the alpha subunit of BK channel family. BK channel protein expression was verified by immunoblotting and confocal microscopy, while the functional role of BK channels in epithelial ion transport mechanisms and cell volume regulation was examined by electrophysiological and morphometric analysis on the intact tissue. BK(Ca) channels appeared to be localized along all the plasma membrane of the enterocytes; the apical part of the villi showed the most intense immunostaining. These channels were silent in basal condition, but were activated on both membranes (apical and basolateral) by increasing intracellular Ca(2+) concentration with the Ca(2+) ionophore ionomycin (1 microM). BK(Ca) channels were also activated on both membranes by hypotonic swelling of the epithelium and their inhibition by 100 nM iberiotoxin (specific BK(Ca) inhibitor) abolished the Regulatory Volume Decrease (RVD) of the intestinal cells after hypotonic swelling. In conclusion, our results demonstrated the molecular and functional expression of high conductance Ca(2+) -activated K(+) channels in eel intestine; the physiological role of these channels is mainly related to the RVD response of the epithelial cells following hypotonic swelling.
Lionetto
Maria G
MG
Department of Biological and Environmental Sciences and Technologies, University of Salento, Leece, Italy. giulia.lionetto@unile.it
Rizzello
Antonia
A
Giordano
Maria E
ME
Maffia
Michele
M
De Nuccio
Francesco
F
Nicolardi
Giuseppe
G
Hoffmann
Else K
EK
Schettino
Trifone
T
D000595:Amino Acid Sequence; D000818:Animals; D004524:Eels / Q000378:metabolism*; D006801:Humans; D007413:Intestinal Mucosa / Q000737:chemistry / Q000378:metabolism*; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D024681:Potassium Channels, Calcium-Activated / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D024681:Potassium Channels, Calcium-Activated