QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online August 18, 2005
Journal of Experimental Biology 208, 3263-3273 (2005)
Published by The Company of Biologists 2005
doi: 10.1242/jeb.01739

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by del Pilar Corena, M. Articles by Linser, P. J. Search for Related Content PubMed PubMed Citation Articles by del Pilar Corena, M. Articles by Linser, P. J.

Carbonic anhydrase in the adult mosquito midgut

Maria del Pilar Corena¹, Leslie VanEkeris¹, Ma. Isabel Salazar², Doria Bowers³, Molly M. Fiedler⁴, David Silverman⁵, Chingkuang Tu⁵ and Paul J. Linser^1,*

¹ The Whitney Laboratory, University of Florida, Saint Augustine, FL 32080, USA
² Arthropod-Borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523, USA
³ Department of Biochemistry, University of North Florida, Jacksonville, FL 32224, USA
⁴ Department of Biochemistry Cell and Molecular Biology, Drake University, Des Moines, IA 50311, USA
⁵ Department of Pharmacology and Therapeutics, University of Florida, Gainesville, FL 32611, USA

View larger version (77K): [in a new window]   Fig. 1. The pH inside the adult Ae. aegypti mosquito midgut. Adult females were fed a mixture containing 0.02% pH indicator. (A) Picture of the dissected mosquito midguts under a stereoscopic microscope. (B) Ae. aegypti females immediately after feeding on the indicator mixture. (C) Dissected midguts of engorged mosquitoes photographed at 10x. (D) pH standards obtained using Cresol Red, Phenol Red, Neutral Red and Thymol Blue. Our results indicate that the pH inside the posterior midgut in adult females from Ae. aegypti ranges between 8.5 and 9.5.

View larger version (82K): [in a new window]   Fig. 2. pH measurements inside the posterior midgut of adult Ae. aegypti, An. gambiae and Cx. tarsalis females indicate that the range is between 8.5 to 9.5. From left to right the columns show the three species, respectively, and the pH standards obtained using Cresol Red, Neutral Red and Thymol Blue.

View larger version (56K): [in a new window]   Fig. 3. CA inhibitors decrease the pH inside the posterior midgut in adult Ae. aegypti females. (A) Cresol Red added to mosquitoes treated with 10–4 mol l–1 MTZ revealed a decrease in pH (left) when compared to those not treated (right). (B) Thymol Blue added to mosquitoes treated with 10–4 mol l–1 MTZ revealed a decrease in pH (left) when compared to those not treated (right). However the effect was not as evident as observed with Cresol Red. Results shown are representative of 10 treated and 10 untreated mosquitoes from 2–4 independent experiments.

View larger version (57K): [in a new window]   Fig. 4. Treatment of Ae. aegypti adult female posterior midgut with MTZ and ACZ combined (10–4 mol l–1 each) resulted in a significant decrease of pH inside the midgut (left) when compared with those midguts dissected from mosquitoes not treated with these inhibitors (right), when compared with the Cresol Red standards (bottom). (A) Dissected midgets. (B) Intact mosquitoes. (C) Dissected midguts photographed at 10x. Results shown are representative of 10–20 treated and untreated individuals from triplicate experiments.

View larger version (69K): [in a new window]   Fig. 5. (A) Grey TIF-8 bit pictures from Ae. aegypti midguts were used for image analysis. The top midgut was treated with 10–4 mol l–1 MTZ and Cresol Red. The lower midgut was treated with Cresol Red in the absence of MTZ. (B) Histogram of average intensity values for treated (10–4 mol l–1) and untreated midgut images (from A) analyzed with ImageQuant TL software from Amersham Biosciences®. Differences were significant (P value<0.0001). 20 specimens (10 treated and 10 untreated) were analyzed in triplicate.

View larger version (76K): [in a new window]   Fig. 6. Hansson's histochemical stain of isolated adult female mosquito midguts. Histochemistry indicates that the enzyme is localized in the midgut of all the species with various degrees of darkening. The enzyme also appears to be associated preferentially with the posterior midgut (black arrows) in several of these species when compared to the anterior midgut (red arrows). (A) Cx. nigripalpus, (B) An. quadrimaculatus, (C) Ae. albopictus, (D) Cx. quinquefasciatus, (E) Ae. aegypti, (F) Oc. taeniorhynchus. Scale bars, 500 µm.

View larger version (73K): [in a new window]   Fig. 7. Hansson's histochemical stain of isolated adult male mosquito midguts indicate that the enzyme is localized in the midgut of all the species with the exception of Oc. taeniorhynchus. (A) An. quadrimaculatus, (B) Ae. albopictus, (C) Cx. quinquefasciatus, (D) Ae. aegypti, (E) An. albimanus, (F) Oc. taeniorhynchus. Black arrows, posterior midgut; red arrows, anterior midgut. Scale bars, 500 µm.

View larger version (11K): [in a new window]   Fig. 8. (A–C) CA activity (as a percentage of total protein) measured in the different regions of the midgut in adult males and females from the species tested revealed that the enzyme is preferentially associated with the posterior midgut in all species, in both males and females. CA activity was detected in anterior (A) and posterior (B) midgut in females and only in the posterior midgut of males (C). (A) CA activity in the anterior midgut of female adult mosquitoes was detected only in Ae. aegypti (highest), Ae. albopictus and Cx. nigripalpus females. (B) CA activity in the female posterior midgut was detected in all species to various degrees. (C) CA activity was detected in the posterior midgut of all species of males tested and was not detected in the anterior midgut of males from any species.