N,N,N’,N’-tetrakis-(2-Pyridylmethyl) ethylenediamine (TPEN) is a heavy metal chelator with high affinity for zinc. TPEN causes important responses in mammalian eggs. For example, these eggs are arrested at the MII stage by the Endogenous Mitotic Inhibitor 2 (Emi2), which prevents activation of the Anaphase Promoting Complex (APC) and degradation of Cyclin B. By chelating zinc, TPEN inactivates Emi2 and eggs undergo spontaneous exit of meiosis and egg activation. TPEN chelates Ca2+ with lower affinity, although in the Endoplasmic Reticulum (ER), where Ca2+ concentrations are high, TPEN may sequester Ca2+ preventing release into the cytoplasm. Initial exposure of TPEN to MII eggs failed to cause spontaneous intracellular Ca2+ release from the ER. Interestingly, in the case of GV oocytes, addition of TPEN caused Ca2+ influx. This influx could be blocked via the addition of 2-APB, a plasma membrane Ca2+ channel blocker. To determine the possible role of TPEN on chelation of ER Ca2+, MII and GV cells were incubated in TPEN and ER Ca2+ released was by exposure to Cyclopiazonic Acid (CPA), a sarco/endoplasmic reticulum (SERCA) pump inhibitor, or Ionomycin (IO), a Ca2+ ionophore. In MII oocytes, the amplitude of the rises caused by CPA and IO, in TPEN-treated oocytes, was smaller than controls and experienced a delay in return to baseline. In GV oocytes, TPEN enhanced rather than reduced Ca2+ responses to CPA and IO. Given its inability to fully chelate ER Ca2+, the use of TPEN as a tool to study Ca2+ homeostasis in mouse oocytes needs additional investigation.
| Identifer | oai:union.ndltd.org:UMASS/oai:scholarworks.umass.edu:theses-2097 |
| Date | 01 January 2013 |
| Creators | Agreda Mccaughin, Robert A |
| Publisher | ScholarWorks@UMass Amherst |
| Source Sets | University of Massachusetts, Amherst |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Masters Theses 1911 - February 2014 |
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