Article

Absence of an intracellular pH change following fertilisation of the mouse egg

Douglas Kline^a1 c1 and James A. Zagray^a1

^a1 Department of Biological Sciences, Kent State University, Kent, Ohio, USA.

Summary

The intracellular pH of the mouse egg was measured during fertilisation to determine whether an increase in pH accompanies activation of this mammalian egg. The pH-sensitive dye BCECF [2′,7′-bis-(2-carboxyethyl)-5(and-6)carboxyfluorescein] was introduced into the mouse egg by incubation in BCECF-AM or by microinjection of dextran-conjugated BCECF. The cells were also loaded with the DNA-specific fluorochrome Hoechst 33342 to confirm fertilisation by observation of Hoechst-stained, decondensing sperm heads in the cytoplasm. The ratio of emission intensities for the dye (494/440 nm excitation wavelengths) was monitored continuously with a photon-counting photomultiplier tube. There was no change in pH during or after fertilisation. Control eggs displayed the expected increase in pH when exposed to NH₄C1. In other experiments, intracellular pH and intracellular Ca²⁺ were monitored simultaneously during fertilisation. The eggs were injected with BCECF dextran and Fura dextran. Fluorescence emission was recorded at excitation wavelengths of 495 nm (BCECF, pH-sensitive wavelength) and 385 nm (Fura, Ca²⁺-sensitive wavelength). A decrease in emission intensity at 385 nm excitation clearly marked the repetitive Ca²⁺ transients at egg activation. There was no change in the fluorescence emitted at 495 nm excitation, indicating an absence of any change in intracellular pH. These results indicate that intracellular alkalinisation of the cytoplasm does not accompany activation of this vertebrate egg.

(Received September 11 1995)

(Accepted September 22 1995)

Key words

• Activation;
• Egg;
• Fertilisation;
• Mouse;
• pH

Correspondence:

^c1 Douglas Kline, Department of Biological Sciences, Kent State University, Kent, OH 44242, USA. Telephone: 216 672-3810. Fax: 216 672-3713. e-mail: dkline@kentvm.kent.edu.