Kalule-Sabiti, Jannat Margaret (1985) A study on the effects of various inhibitors on the ATPase activity in, and fluid secretion by Malpighian tubules of Locusta Migratoria L. Doctoral thesis, Durham University.
A study of the effects of some inhibitors on both the activities of ATPases isolated from Locusta Malpighian tubules microsomal preparations and in vitro Malpighian tubule fluid production has been carried out. Inhibitors investigated included ouabain, vanadate, furosemide, acetazolamide, SITS and SCN(^_). It was important to establish the effects of temperature and[K(^+)] on the inhibitory action of both ouabain and vanadate. Ouabaindid not significantly inhibit either Na(^+), K(^+)-ATPase or in vitro Malpighian fluid production at temperatures below 30 C. In contrast vanadate was negatively influenced by temperature. Vanadate was most effective at temperatures between 20 and 30 C. High [K ]+ +enhanced vanadate but antagonised ouabain inhibition of Na , K -ATPase. Under optimal conditions ouabain inhibited both Na(^+), K(+)-ATPase (pI(_50) = 6.8) and in vitro Malpighian tubule fluid production (piso = 4.3). Similarly vanadate inhibited Na*, K*-ATPase (pI(_50) = 5.8) and in vitro Malpighian tubule fluid secretion (pI(_50) = 4.8). Vanadate was found to be nonspecific as it also inhibited both Mg(^2+),-ATPase and Mg(^2+), HCO(^-)(_3) -ATPase activities. With the exception of SITS, all other inhibitors studied inhibited in vitro Malpighian tubule fluid section. However, SITS was found to be a strong inhibitor of Mg(^2+), HCO(^-)(_3) -ATPase. All results are discussed with reference to the role of ATPases in the process of ion and water transport across the Malpighian tubules. Cytochemical and biochemical studies based on ERNST (1972a,b) technique were carried out as the first attempt to localise Na(^+),K(^+)-ATPase in Locusta Malpighian tubule cells. K(^+)-NPPase was localised mainly along the cytoplasmic side of the basal cell membrane in foldings but was inconsistently inhibited by ouabain. Biochemical studies showed that paraformaldehyde fixation of the Malpighian tubules and inclusion of 20mM Sr C1(_2) in the standard incubation medium reduced the total NPPase activity by 73%. Use of β-glycerophosphate indicated that the reaction product observed in the present study was not due to nonspecific alkaline phosphatase activity. However, the results were inconclusive and led to further questioning of the validity of ERNST (1972b) procedure in localisation of Na(^+),K(^+)-ATPase in insect epithelia.
|Item Type:||Thesis (Doctoral)|
|Award:||Doctor of Philosophy|
|Copyright:||Copyright of this thesis is held by the author|
|Deposited On:||16 Jul 2013 10:57|