Localization and vasopressin regulation of the Na⁺-K⁺-2Cl⁻ cotransporter in the distal colonic epithelium.
OBJECTIVE
To investigate whether Na(+)-K(+)-2Cl(-) cotransporter (NKCC2) is expressed in the mouse distal colonic epithelia and whether it is regulated by vasopressin in the colon.


METHODS
The mRNA expression of NKCC2 in the mouse colonic mucosa was examined by reverse transcription-polymerase chain reaction. NKCC trafficking in the colon stimulated by 1-D-amino(8-D-arginine)-vasopressin (dDAVP) infusion (10 ng/mouse, intraperitoneal injection ) within 15 min, 30 min and 1h was investigated by laser confocal scanning microscopy. Total and membrane NKCC2 expression in the colonic mucosa from control and dDAVP-treated mice was detected by Western blotting. Short circuit current method was performed to determine regulation of NKCC2 by vasopressin in the colon.


RESULTS
NKCC2 was predominantly located in the apical region of the surface of the distal colonic epithelia; by comparison, a large amount of NKCC1 was distributed in the basolateral membrane of the lower crypt epithelia of the mouse distal colon. Short-term treatment with dDAVP, a V2-type receptor-specific vasopressin analog, induced NKCC2 re-distribution, i.e., NKCC2 traffics to the apical membrane after dDAVP stimulation. In contrast, no obvious NKCC1 membrane translocation was observed. Western blotting results confirmed that membrane NKCC2 had significantly higher abundance in the dDAVP-treated mouse colonic mucosa relative to that in the untreated control, which is consistent with our immunostaining data. Moreover, the short-circuit current method combined with a NKCC2 inhibitor demonstrated that NKCC2 was also activated by serosal vasopressin in isolated distal colonic mucosa.


CONCLUSIONS
Our results provide direct evidence that vasopressin also plays an important role in the colonic epithelia by stimulating NKCC2 trafficking to the apical membrane and inducing NKCC2-mediated ion transport.
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Hong
H
Xue
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Zi-Juan
ZJ
Zhang
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Xiao-Shuang
XS
Li
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Hai-Mei
HM
Sun
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Qian
Q
Kang
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Bo
B
Wu
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Ya-Xi
YX
Wang
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
Wan-Jing
WJ
Zou
Hong Xue, Xiao-Shuang Li, Hai-Mei Sun, Qian Kang, Bo Wu, Ya-Xi Wang, Wan-Jing Zou, De-Shan Zhou, Department of Histology and Embryology, School of Basic Medical Science, Capital Medical University, Beijing 100069, China.
De-Shan
DS
Zhou
D000818:Animals; D003106:Colon / Q000187:drug effects* / Q000378:metabolism; D003894:Deamino Arginine Vasopressin / Q000008:administration & dosage / Q000494:pharmacology*; D007263:Infusions, Parenteral; D007413:Intestinal Mucosa / Q000187:drug effects* / Q000378:metabolism; D017136:Ion Transport; D008297:Male; D008564:Membrane Potentials; D008810:Mice, Inbred C57BL; D021381:Protein Transport; D012333:RNA, Messenger / Q000378:metabolism; D064466:Solute Carrier Family 12, Member 1 / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D012333:RNA, Messenger; C578715:Slc12a1 protein, mouse; D064466:Solute Carrier Family 12, Member 1; D003894:Deamino Arginine Vasopressin
Apical membrane; Distal colonic epithelia; Na+-K+-2Cl- cotransporter; Trafficking; Vasopressin