In Chapter 1 we characterized the frequency, amplitude and duration of (Ca$\sp{2+}\rbrack\sb{\rm i}$ rises in fertilized rabbit eggs loaded with fura-2 dextran. Their amplitude decreased and duration increased as fertilization progressed. Injection of 1,4,5 inositol trisphosphate (InsP3; IICR) or addition of thimerosal elicited (Ca$\sp{2+}\rbrack\sb{\rm i}$ rises which were blocked by heparin, an InsP3 receptor antagonist. Ryanodine did not evoke Ca$\sp{2+}$ release. These results indicate that IICR is likely stimulated during fertilization. Fertilization (Ca$\sp{2+}\rbrack\sb{\rm i}$ changes were examined in in vitro matured bovine eggs (Chapter 2). Fertilized eggs exhibited different intervals between (Ca$\sp{2+}\rbrack\sb{\rm i}$ rises which ranged from 15 min to more than 60 min. Unfertilized eggs were responsive to InsP3 injection and thimerosal exposure, although the frequency of the (Ca$\sp{2+}\rbrack\sb{\rm i}$ responses was shorter than the periodicity observed during fertilization. The mechanisms that generate (Ca$\sp{2+}\rbrack\sb{\rm i}$ oscillations were examined in fertilized rabbit eggs (Chapter 3). Heparin blocked or delayed (Ca$\sp{2+}\rbrack\sb{\rm i}$ oscillations. Oscillating eggs exhibited Ca$\sp{2+}$ release in response to CaCl2 injection. In unfertilized eggs, injection of GTP (S) induced (Ca$\sp{2+}\rbrack\sb{\rm i}$ oscillations and enhanced the response to CaCl2 injection. Injection of InsP3 or CaCl2 elicited full Ca$\sp{2+}$ responses that reset the periodicity of ensuing oscillations. Thus, IICR participates in the generation of (Ca$\sp{2+}\rbrack\sb{\rm i}$ oscillations and the unloading of the stores does not explain the interval between (Ca$\sp{2+}\rbrack\sb{\rm i}$ rises. The signaling pathways possibly stimulated by the sperm during fertilization were investigated in unfertilized bovine eggs (Chapter 4). Injection of GTP (S) or InsP3 evoked (Ca$\sp{2+}\rbrack\sb{\rm i}$ oscillations. Preinjection of heparin blocked sperm-mediated egg activation. Thimerosal elicited (Ca$\sp{2+}\rbrack\sb{\rm i}$ oscillations which were not inhibited by heparin or ryanodine. The data suggest that bovine eggs possess a GTP-linked phosphoinositide pathway which appears to be stimulated by the sperm during fertilization. In Chapter 5 the amplitude and duration of (Ca$\sp{2+}\rbrack\sb{\rm i}$ rises elicited by electrical pulses of different strengths and duration administered in variable extracellular Ca$\sp{2+}$ concentrations was reported. As these parameters increased, so did the peak (Ca$\sp{2+}\rbrack\sb{\rm i}$ elicited by the pulse. Young and aged eggs exhibited similar (Ca$\sp{2+}\rbrack\sb{\rm i}$ rises when stimulated with identical pulses. However, their activation rates were different. Thus, aged eggs are more sensitive to a given (Ca$\sp{2+}\rbrack\sb{\rm i}$ rise.
| Identifer | oai:union.ndltd.org:UMASS/oai:scholarworks.umass.edu:dissertations-8745 |
| Date | 01 January 1993 |
| Creators | Fissore, Rafael Antonio |
| Publisher | ScholarWorks@UMass Amherst |
| Source Sets | University of Massachusetts, Amherst |
| Language | English |
| Detected Language | English |
| Type | text |
| Source | Doctoral Dissertations Available from Proquest |
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