Document Detail


Stimulation of repetitive calcium transients in mouse eggs.
MedLine Citation:
PMID:  7650607     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
1. We have combined cell membrane electroporation by electrical field (EF) stimulation with a rapid perfusion system in order to stimulate repetitive increases in cytoplasmic free [Ca2+] ([Ca2+]i) in mouse eggs. [Ca2+]i was monitored by ratio fluorescent measurements of intracellular indo-1 on individual eggs. The conditions required to cause different types of [Ca2+]i increases were established and the effects of these [Ca2+]i changes upon egg activation examined. 2. The rapid perfusion of non-ionic medium caused a single [Ca2+]i increase. However, to generate repetitive [Ca2+]i increases, eggs were exposed to EF pulses in the presence of Ca2+ and then washed rapidly with culture medium. Sequential EF pulse application led to prolonged elevation of [Ca2+]i levels and eventual cell lysis unless rapid reperfusion with culture medium was achieved. Transient increases in [Ca2+]i in eggs could also be generated by EF pulses in the presence of inositol 1,4,5-trisphosphate (InsP3). 3. In response to EF stimulation fertilized eggs showed [Ca2+]i increases that were enhanced relative to unfertilized eggs. The responses in these fertilized eggs were often followed by repetitive [Ca2+]i oscillations, despite the fact that the [Ca2+]i oscillations associated with sperm penetration had ceased by this stage. 4. In unfertilized mouse eggs the [Ca2+]i increases appeared to be due to direct cation influx since repeated EF pulses caused repeated influx of Mn2+ as monitored by quenching of fluorescence of fura-2 loaded eggs. 5. Under conditions that stimulated reproducible patterns of [Ca2+]i transients we found that a single large [Ca2+]i transient did not cause significant egg activation, but that inducing repetitive [Ca2+]i transients was effective in activating eggs. The speed of activation as judged by the rate of pronuclear formation was also dependent upon the frequency of pulse application. 6. These data show that combining EF pulses with a rapid and precise sequential perfusion system can be used to manipulate [Ca2+]i levels in mammalian eggs. This provides a means of artificial mimicry of the [Ca2+]i transients seen after fertilization. It appears that Ca2+ influx during EF pulses does not cause significant Ca2+ release from internal stores in unfertilized eggs, but after fertilization Ca2+ influx does induce Ca2+ release. It is also apparent that mouse eggs are more successfully activated by repetitive [Ca2+]i increases than by single large [Ca2+]i rises. We suggest that our data provide direct evidence for the hypothesis that a cellular response to oscillations of intracellular [Ca2+]i can be distinct from that to monotonic rises in [Ca2+]i.
Authors:
J P Ozil; K Swann
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  The Journal of physiology     Volume:  483 ( Pt 2)     ISSN:  0022-3751     ISO Abbreviation:  J. Physiol. (Lond.)     Publication Date:  1995 Mar 
Date Detail:
Created Date:  1995-09-28     Completed Date:  1995-09-28     Revised Date:  2009-11-18    
Medline Journal Info:
Nlm Unique ID:  0266262     Medline TA:  J Physiol     Country:  ENGLAND    
Other Details:
Languages:  eng     Pagination:  331-46     Citation Subset:  IM    
Affiliation:
Institut National de la Recherche Agronomique, Unité de Biologie de la Fécondation, Jouy en Josas, France.
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MeSH Terms
Descriptor/Qualifier:
Animals
Calcium / metabolism*,  pharmacology
Culture Media
Electric Conductivity
Female
Fluorescent Dyes
Indoles
Inositol 1,4,5-Trisphosphate / pharmacology
Mice
Mice, Inbred C57BL
Mice, Inbred CBA
Ovum / chemistry,  metabolism*
Parthenogenesis / physiology
Periodicity
Time Factors
Zygote / physiology
Chemical
Reg. No./Substance:
0/Culture Media; 0/Fluorescent Dyes; 0/Indoles; 7440-70-2/Calcium; 85166-31-0/Inositol 1,4,5-Trisphosphate; 96314-96-4/indo-1
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