Browsing by Author "Parkel, Sven"

Filter results by typing the first few letters
Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    [3H]WAY 100635 sidumine roti hipokampuse membraanidele
    (2004) Parkel, Sven; Rinken, Ago, juhendaja
  • Thumbnail Image
    Item
    Ligand binding to 5-HT1A receptors and its regulation by Mg2+ and Mn2+
    (Tartu University Press, 2010-01-14T08:04:14Z) Parkel, Sven
    In the first part of this thesis the binding of 5-HT1A receptor specific antagonist [3H]WAY100635 to rat brain membranes was characterized. It was found, that the binding on lower concentrations was considerably slow, and a notable amount of receptors will be inactivated before thermodynamic equilibrium can be reached, yet on higher concentrations rapid increase in reaction rate indicative of cooperative binding was detected. This means that [3H]WAY100635 is an excellent tool for determining receptor number in a given sample, but not a very good ligand in equilibrium binding studies. In the second part of this thesis the influence of Mn2+ in comparison to Mg2+ to signal transduction via 5-HT1A receptors was measured. It was shown that Mn2+ can stabilize greater number of agonist high-affinity binding sites, while the size of this effect depended on the used brain region (different in hippocampal and cortical membranes) and also on the GTPγS concentration. Since GTPγS activates G-proteins, sensitivity to its concentration refers to the involvement of G-proteins in the enhancing regulation by Mn2+. Sf9 cell lines were created for comparison expressing only 5-HT1A receptor or the receptor with Gi or Gs protein. Agonist high affinity binding was achieved only in cell line expressing the receptor with Gi protein, and the sensitivity of this cell line to Mn2+ was similar to cortical preparations but different from hippocampal membranes. To clarify the role of G-proteins on this regulation, influence of Mn2+ compared to Mg2+ to the affinity of guanosine nucleotides (GDP and GTPγS) binding to G-proteins was measured. Mn2+ lowered the affinity of both GDP and GTPγS compared to Mg2+, and the size of the effect was again dependent on used brain region, showing that there are differences in the composition of signaling complexes that are formed with 5-HT1A receptor in different brain regions. Additionally there seem to be differences in the sensitivity of these complexes to Mn2+-ions.