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Vacuolar-ATPase (V-ATPase) Mediates Progesterone-Induced Uterine Fluid Acidification in Rats

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Abstract

We hypothesized that progesterone-induced decrease in uterine fluid pH involves V-ATPase. In this study, expression and functional activity of V-ATPase in uterus were investigated under progesterone influence. Ovariectomized adult female rats received subcutaneous injection of estradiol-17β (1 µg/kg/day) or progesterone (20 mg/kg/day) for 3 days or 3 days estradiol-17β followed by 3 days vehicle, progesterone, or estradiol-17β plus progesterone. Mifepristone, a progesterone receptor blocker, was concomitantly given to the rats which received progesterone. A day after last injection, rate of uterine fluid secretion, its HCO3 − concentration, and pH were determined via in vivo uterine perfusion in rats under anesthesia. V-ATPase inhibitor, bafilomycin, was introduced into the perfusion buffer, and changes in these parameters were observed. Expression of V-ATPase A1 and B1/2 proteins and mRNAs in uterus were quantified by Western blotting and real-time PCR, respectively. Distribution of these proteins was observed by immunohistochemistry. Our findings showed that under progesterone influence, uterine fluid secretion rate, HCO3 − concentration, and pH were significantly reduced. Administration of bafilomycin did not cause significant changes in fluid secretion rate; however, HCO3 − concentration and pH were significantly elevated. In parallel with these changes, expression of V-ATPase A1 and B1/2 proteins and mRNAs were significantly increased with these proteins highly distributed in uterine luminal and glandular epithelia. In conclusion, increased expression and functional activity of V-ATPase were most likely responsible for the decreased in uterine fluid pH observed under progesterone influence.

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Abbreviations

V:

Vacuolar

HCO3 − :

Bicarbonate

mRNA:

Messenger RNA

CFTR:

Cystic fibrosis transmembrane regulator

a2V:

a2V-ATPase

LIF:

Leukemia inhibitory factor

Il1β:

Interleukins

IACUC:

Institutional Animal Care and Use Committee

RIA:

Radioimmunoassay

Na2HCO3 :

Sodium bicarbonate

Na2HPO4 :

Sodium phosphate dibasic

MgSO4 :

Magnesium sulfate

HEPES:

4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid

CaCl2 :

Calcium chloride

PEPC:

Phosphoenolpyruvate carboxylase

MDH:

Malate dehydrogenase

H2O2 :

Hydrogen peroxide

PBS:

Phosphate-buffered saline

DAB:

3,3′-Diaminobenzidine

PVDF:

Polyvinyilidenedifluoride

GAPDH:

Glyceraldehyde 3-phosphate dehydrogenase

UNG:

Uracil N-glycosylase

NHE:

Sodium hydrogen exchanger

ENaC:

Epithelial sodium channel

PGs:

Prostaglandins

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Authors and Affiliations

  1. Department of Physiology, Faculty of Medicine, University of Malaya, 50603, Kuala Lumpur, Malaysia

    Kamarulzaman Karim, Nelli Giribabu & Naguib Salleh

  2. Department of Molecular Medicine, Faculty of Medicine, University of Malaya, 50603, Kuala Lumpur, Malaysia

    Sekaran Muniandy

Authors
  1. Kamarulzaman Karim
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  2. Nelli Giribabu
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  3. Sekaran Muniandy
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  4. Naguib Salleh
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Corresponding author

Correspondence to Naguib Salleh.

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Conflict of Interest

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Funding

This study was funded by PPP Grant (PV069-2014A), University of Malaya, Kuala Lumpur, Malaysia.

Additional information

This work was done in Department of Physiology and Department of Molecular Medicine, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.

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Karim, K., Giribabu, N., Muniandy, S. et al. Vacuolar-ATPase (V-ATPase) Mediates Progesterone-Induced Uterine Fluid Acidification in Rats. J Membrane Biol 249, 65–76 (2016). https://doi.org/10.1007/s00232-015-9848-z

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  • DOI: https://doi.org/10.1007/s00232-015-9848-z

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