Adult body weight is programmed by a redox-regulated and energy-dependent process during the pronuclear stage in mouse

Abstract

In mammals fertilization triggers a series of Ca(2+) oscillations that not only are essential for events of egg activation but also stimulate oxidative phosphorylation. Little is known, however, about the relationship between quantitative changes in egg metabolism and specific long-term effects in offspring. This study assessed whether post-natal growth is modulated by early transient changes in NAD(P)H and FAD(2+) in zygotes. We report that experimentally manipulating the redox potential of fertilized eggs during the pronuclear (PN) stage affects post-natal body weight. Exogenous pyruvate induces NAD(P)H oxidation and stimulates mitochondrial activity with resulting offspring that are persistently and significantly smaller than controls. Exogenous lactate stimulates NAD(+) reduction and impairs mitochondrial activity, and produces offspring that are smaller than controls at weaning but catch up after weaning. Cytosolic alkalization increases NAD(P)(+) reduction and offspring of normal birth-weight become significantly and persistently larger than controls. These results constitute the first report that post-natal growth rate is ultimately linked to modulation of NAD(P)H and FAD(2+) concentration as early as the PN stage.

Figure 1. Energy starvation and developmental responses.

Metabolic profiles and post-natal phenotypes induced by “starvation.” (A) Time course of intracellular NAD(P)H (blue), FAD²⁺ (red) signals (grey levels, g.l., 14 bits) and pH_i fluorescence (green) of eggs subjected to M_starv. The initial NAD(P)H biphasic curve is due to the high lactate/pyruvate ratio in M16 that delays intracellular depletion of carbohydrates. (B) Growth profiles of control (black lines) and experimental animals (dark pink lines). The data are expressed as mean ± SEM. (C) Standardized plots of the total Z values (n = 210). The upper limits of the shaded zones (grey for controls and dark pink for the experimental values) are the normalized weight profiles of males issued from the larger litter sizes (8, 7 and 6) and the lower limits, from the smaller litter sizes (5, 4, 3 and 2). (D) Relative weight gains (RWG) of experimental animals according to the larger or smaller litter size group. In this figure and in figures 2–4, the stars in panels B and C denote significant differences between average experimental and control values (with a P-value at least <0.04) when compared to controls.

Figure 2. Exogenous pyruvate and developmental responses.

Metabolic profiles and post-natal phenotypes induced by “pyruvate.” (A) Time course of intracellular NAD(P)H (blue), FAD²⁺ (red) signal (grey levels, g.l., 14 bits) and pH_i fluorescence (green) of eggs subjected to M_pyr. (B) Growth profiles of control (black lines) and experimental (red lines) animals. The growth profiles of M_starv animals from Fig. 1B are plotted as dark pink lines for comparison. The data are expressed as mean ± SEM. (C) Standardized plots of total Z values (n = 210). The upper limits of the shaded zones (grey for the controls and red for the experimental values) are the normalized weight profiles of males issued from the larger litter sizes (8, 7 and 6) and the lower limits, from the smaller litter sizes (5, 4, 3 and 2). (D) Relative weight gains (RWG) of experimental animals according to the larger or smaller litter size group.

Figure 3. Exogenous lactate and developmental responses.

Metabolic profiles and post-natal phenotypes induced by “Lactate.” (A) Time course of intracellular NAD(P)H (blue), FAD²⁺ (red) signals (grey levels, g.l., 14 bits) and pH_i fluorescence (green) of eggs subjected to M_lac. (B) Growth profiles of control and experimental animals. The growth profiles of M_pyr animals from Fig. 2B are plotted as red lines for comparison. The blue crosses and plus symbols show when the M_lac profiles are significantly higher than the M_pyr profiles for males and females, respectively. The data are expressed as mean ± SEM. (C) Standardized plots of total Z values (n = 210). The upper limits of the shaded zones (grey for the controls and blue for the experimental values) are the normalized weigh profiles of males issued from the larger litter sizes (8, 7 and 6) and the lower limits from the smaller litter sizes (5, 4, 3 and 2). (D) Relative weight gains (RWG) of experimental animals according to the larger or smaller litter size group.

Figure 4. Alkalization and developmental responses.

Metabolic profiles and post-natal phenotypes induced by “alkalization.” (A) Time course of intracellular NAD(P)H (blue), FAD²⁺ (red) signals (grey levels, g.l., 14 bits) and pH_i fluorescence (green) of eggs subjected to M_pH. (B) Growth profiles of control and experimental animals. The growth profiles of M_pyr animals from Fig. 2B are plotted as red lines for comparison. The data are expressed as mean ± SEM. (C) Standardized plots of total Z values (n = 210). The upper limits of the shaded zones (grey for the controls and green for the experimental values) are the normalized weight profiles of males issued from the larger litter sizes (8, 7 and 6) and the lower limits from the smaller litter sizes (5, 4, 3 and 2). (D) Relative weight gains (RWG) of M_pH experimental animals according to the larger or smaller litter size group (green lines). The RWG of M_pyr animals from Fig. 2B are plotted as red lines for comparison.

Figure 5. Post-natal outcomes resulting from zygotic metabolic sensor activities.

The diagram represents the linkages between the composition of the culture media, the NAD(P)H and FAD²⁺ concentrations and the adult phenotype, that are mediated by a zygotic metabolic sensor.