Amphibian oocytes have been extensively used for heterologous expression of membrane proteins for studying their biochemical and biophysical properties. So far, Xenopus laevis is the main amphibian used as oocytes source to express aquaglyceroporins in order to assess water and solutes permeability. However, this well-established amphibian model represents a threat to the biodiversity in many countries, especially in those from tropical regions. For that reason, the import of Xenopus laevis is subjected to strict control, which essentially has restricted its use in these regions. Therefore, a wider variety of expression systems for aquaglyceroporins is needed. Rhinella marina is extensively distributed in the Americas and its native range spreads from South America to Texas, US. Here we report the use of Rhinella marina oocytes as an alternative expression system for aquaglyceroporins and demonstrated its suitability to determine the permeability to water and non-ionic solutes. Rhinella marina oocytes were able to functionally express channels from human and the protozoan pathogen Trypanosoma brucei, two very distant organisms on the evolutionary scale. Permeability values obtained from Rhinella marina oocytes expressing members of aquaporin family were similar and comparable to those values reported in the literature for the same channels expressed in Xenopus laevis oocytes.