Final Draft Lab 4

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Effects of oligomycin, ouabain, p-Aminohippurate, sodium concentrations and potassium concentrations on active transport of the Malpighian tubules of Acheta domesticus Key Words: Malpighian tubules, Acheta domesticus, PAH, ouabain, oligomycin, Na+, K+

Silvia Kilic
100488712
BIOL 3040U
CRN: 70041
Thursday, March 5th, 2015.
Results
The Malpighian tubules were incubated in the solutions for 15 minutes before being observed. The transport of CPR into the lumen was highest when there was no addition of oligomycin, ouabain and p-aminohippurate (PAH). The least transport occurred from the addition of ouabain and PAH (table 1). There was a strong positive correlation (R2=0.9881) between increased potassium concentration in the CPR solution and the transport of the solution into the tubule (figure 1). The least tubule transport occurred at 0mM potassium (score of 0) and the most tubule transport occurred at 10mM potassium (score of 4) (figure 1). There was a strong positive correlation (R2=0.9871) between increased sodium concentration in the CPR solution and the transport of the solution into the tubule (figure 2). The least tubule transport occurred at 0mM sodium (score of 0.083 ± 0.204) and the most tubule transport occurred at 160mM sodium (score of 3.66 ± 0.816) (figure 2). Discussion

The experiment exemplified the effects of oligomycin, p-Aminohippurate (PAH), sodium and potassium on the active transport of chlorophenol red (CPR) into the cricket, Acheta domesticus, Malpighian tubules (MT). The study of the active transport system in crickets is useful in modeling how active transport works and how it can be affected or inhibited. This information can be transferred to the mammalian kidney that also has active transport present in order to move solutes between the tubules and capillaries. This was a controlled experiment. The CPR solution without any added contents acted as a control. It was kept constant throughout each solution and gave a positive result, transported the CPR solution into the Malpighian tubules. The person doing the assessments was unaware of the incubation solutions used in order to make a fair assessment. If they were aware, a biased opinion would most likely be made if there are expected results. For example, oligomycin was predicted to give a negative result for the transport of CPR, if the assessor was aware of this they may have made a biased opinion stating a value of 0 rather than the given value of 1 (table 1). There does not seem to be a threshold concentration for sodium or potassium. The results indicated that with increased potassium and sodium concentration, the transport of solution into the MT also increased. The transport of CPR with oligomycin into the tubules from the extracellular fluid (ECF) was given a score of 1. This indicates that there was very little transport of dye. Oligomycin blocks the F0 portion of the F0F1-ATP synthase which is fixed in the membrane and transports protons (Senior and Weber, 2003). The proton gradient generated is used to synthesis ATP. Both Na+/K+ATPase and V-type H+ -ATPase need the energy from ATP in order to maintain the sodium/potassium gradient that drives the Na+-dicarboxylate co-transport (Dantzler, 2002). The transport of CPR is dependent on the exchange of a dicarboxylate, which if absent due to the lack of sodium will cause no transport of CPR into the tubules (Dantzler, 2002). PAH was observed to cause no transport of CPR, score of 0 (table 1). Ianowski et al. (2003) explained opposing results, Malpighian tubules are able to transport PAH, and that this transport is dependent upon the Na+ gradient. PAH is a small organic anion which uses diacrboxylate exchange in the same way CPR does, so it should have been able to be transported across the tubules (Masereeuw et al., 2003). The solution containing CPR and ouabian had a score of 0, indicating no transport into the Malpighian tubules. Ouabain inhibits the Na+/K+...
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