卵細胞Ca離子穩定攸關卵子受孕之過程
卵細胞細胞膜上之K+離子ATP閥可控制Ca離子之進出進而影響Ca離子穩定
Pinacidil and glibenclamide可控制K+離子ATP閥，進而維持卵子細胞內Ca離子穩定

Hum Reprod. 2016 Feb;31(2):287-97. doi: 10.1093/humrep/dev300. Epub 2015 Dec 18.

A spontaneous increase in intracellular Ca2+ in metaphase II human oocytes in vitro can be prevented by drugs targeting ATP-sensitive K+ channels.

Fernandes G1, Dasai N1, Kozlova N1, Mojadadi A2, Gall M3, Drew E3, Barratt E3, Madamidola OA4, Brown SG5, Milne AM1, Martins da Silva SJ6, Whalley KM3,Barratt CL6, Jovanović A7.

Author information

Abstract

STUDY QUESTION:

Could drugs targeting ATP-sensitive K(+) (KATP) channels prevent any spontaneous increase in intracellular Ca(2+) that may occur in human metaphase II (MII) oocytes under in vitro conditions?

SUMMARY ANSWER:

Pinacidil, a KATP channel opener, and glibenclamide, a KATP channel blocker, prevent a spontaneous increase in intracellular Ca(2+) in human MII oocytes.

WHAT IS KNOWN ALREADY:

The quality of the oocyte and maintenance of this quality during in vitro processing in the assisted reproductive technology (ART) laboratory is of critical importance to successful embryo development and a healthy live birth. Maintenance of Ca(2+) homeostasis is crucial for cell wellbeing and increased intracellular Ca(2+) levels is a well-established indicator of cell stress.

STUDY DESIGN, SIZE, DURATION:

Supernumerary human oocytes (n = 102) collected during IVF/ICSI treatment that failed to fertilize were used from October 2013 to July 2015. All experiments were performed on mature (MII) oocytes. Dynamics of intracellular Ca(2+) levels were monitored in oocytes in the following experimental groups: (i) Control, (ii) Dimethyl sulfoxide (DMSO; used to dissolve pinacidil, glibenclamide and 2,4-Dinitrophenol (DNP)), (iii) Pinacidil, (iv) Glibenclamide, (v) DNP: an inhibitor of oxidative phosphorylation, (vi) Pinacidil and DNP and (vii) Glibenclamide and DNP.

PARTICIPANTS/MATERIALS/SETTINGS/METHODS:

Oocytes were collected under sedation as part of routine treatment at an assisted conception unit from healthy women (mean ± SD) age 34.1 ± 0.6 years, n = 41. Those surplus to clinical use were donated for research. Oocytes were loaded with Fluo-3 Ca(2+)-sensitive dye, and monitored by laser confocal microscopy for 2 h at 10 min intervals. Time between oocyte collection and start of Ca(2+) monitoring was 80.4 ± 2.1 h.

MAIN RESULTS AND THE ROLE OF CHANCE:

Intracellular levels of Ca(2+) increased under in vitro conditions with no deliberate challenge, as shown by Fluo-3 fluorescence increasing from 61.0 ± 11.8 AU (AU = arbitrary units; n = 23) to 91.8 ± 14.0 AU (n = 19; P < 0.001) after 2 h of monitoring. Pinacidil (100 µM) inhibited this increase in Ca(2+) (85.3 ± 12.3 AU at the beginning of the experiment, 81.7 ± 11.0 AU at the end of the experiment; n = 13; P = 0.616). Glibenclamide (100 µM) also inhibited the increase in Ca(2+) (74.7 ± 10.6 AU at the beginning and 71.8 ± 10.9 AU at the end of the experiment; n = 13; P = 0.851. DNP (100 mM) induced an increase in intracellular Ca(2+) that was inhibited by glibenclamide (100 µM; n = 9) but not by pinacidil (100 µM; n = 5).