The electrical block to polyspermy induced by an intracellular Ca²⁺ increase at fertilization of the clawed frogs, Xenopus laevis and Xenopus tropicalis

Abstract

Polyspermy blocking, to ensure monospermic fertilization, is necessary for normal diploid development in most animals. We have demonstrated here that monospermy in the clawed frog, Xenopus tropicalis, as well as in X. laevis, is ensured by a fast, electrical block to polyspermy on the egg plasma membrane after the entry of the first sperm, which is mediated by the positive‐going fertilization potential. An intracellular Ca2+ concentration ([Ca2+]i) at the sperm entry site was propagated as a Ca2+ wave over the whole egg cytoplasm. In the X. tropicalis eggs fertilized in 10% Steinberg's solution, the positive‐going fertilization potential of +27 mV was generated by opening of Ca2+‐activated Cl−‐channels (CaCCs). The fertilization was completely inhibited when the egg's membrane potential was clamped at +10 mV and 0 mV in X. tropicalis and X. laevis, respectively. In X. tropicalis, a small number of eggs were fertilized at 0 mV. In the eggs whose membrane potential was clamped below −10 mV, a large increase in inward current, the fertilization current, was recorded and allowed polyspermy to occur. A small initial step‐like current (IS current) was observed at the beginning of the increase in the fertilization current. As the IS current was elicited soon after a small increase in [Ca2+]i, this is probably mediated by the opening of CaCCs. This study not only characterized the fast and electrical polyspermy in X. tropicalis, but also explained that the initial phase of [Ca2+]i increase causes IS current during the early phase of egg activation of Xenopus fertilization.

Cellular dynamics of double fertilization and early embryogenesis in flowering plants

“…The intracellular Ca 2+ increase acts as a signal, resulting, for instance, in dephosphorylation of mitogen‐activated protein kinase (MAPK). This inhibits extra sperm–egg fusions and sperm attraction in jellyfish and, in monospermic frogs, induces a reversal of electrical properties between the interior and exterior of the egg membrane, blocking additional sperm entry by activating an efflux of Cl − (Arakawa, Takeda, Tachibana, & Deguchi, 2014; Iwao & Izaki, 2018; Watabe et al, 2019).…”

Physiological polyspermy: Selection of a sperm nucleus for the development of diploid genomes in amphibians

“…After the fast polyspermy block, the cortical granules are discharged to form a fertilization envelope, which prevents the approach of sperm at the egg surface for several minutes after fertilization (Grey, Working, & Hedrick, ; Hedrick, ; Wong & Wessel, ), which is referred to as the slow but permanent block to polyspermy. However, several sperm can enter a monospermic frog egg through artificial reduction of the fertilization potential (Grey, Bastiani, Webb, & Schertel, ; Watabe et al, ) or the insemination of immature eggs whose responses against sperm entry are slower than those of mature eggs (Iwao, ; Schlichter & Elinson, ). In "pathological polyspermy", all incorporated sperm nuclei form sperm pronuclei with well‐developed asters (Figures b and b).…”

“…However, several sperm can enter a monospermic frog egg through artificial reduction of the fertilization potential (Grey, Bastiani, Webb, & Schertel, 1982;Watabe et al, 2019) or the insemination of immature eggs whose responses against sperm entry are slower than those of mature eggs (Iwao, 1987;Schlichter & Elinson, 1981). In "pathological polyspermy", all incorporated sperm nuclei form sperm pronuclei with well-developed asters (Figures 1b and 2b).…”

“…In most animals, only one sperm is incorporated into the egg, which is referred to as monospermy (Figures a and a), and extra sperm is prevented from entering the fertilized egg by several blocks against polyspermy (Iwao & Izaki, ). In monospermic frogs such as the South African clawed frog, Xenopus laevis , and the western clawed frog, Xenopus tropicalis , the membrane potential of unfertilized eggs is negative (~−20 mV), and a positive fertilization potential of 7–27 mV is elicited at sperm entry (Watabe et al, ; Webb & Nuccitelli, ). The fertilization potential is dependent on the opening of Ca 2+ ‐activated Cl − ‐channels and lasts for 10–15 min after fertilization.…”

“…The fertilization potential is dependent on the opening of Ca 2+ ‐activated Cl − ‐channels and lasts for 10–15 min after fertilization. When the membrane voltage of unfertilized eggs remains higher than 0 mV, fertilization is prevented, whereas maintenance of the membrane voltage at less than −10 mV causes polyspermy (Glahn & Nuccitelli, ; Watabe et al, ). Thus, monospermy is ensured by the positive fertilization potential operating as a fast electrical block to polyspermy at the egg plasma membrane (Iwao & Izaki, ).…”