Exploring haemoglobin respiration and oxidation states in avian and mammalian species

Abstract

The native avian species of New Zealand, which evolved in the absence of mammalian predators, are highly susceptible to invasive mammals. Current methods to control these predators often fall short and pose significant risks to non-target birds. Therefore, identifying specific physiological targets for developing species-specific toxicants is crucial. Haemoglobin (Hb) is responsible for oxygen transport within erythrocytes across all vertebrates. It transforms into methaemoglobin (metHb) during oxidation, inhibiting oxygen binding and leading to methaemoglobinaemia. Unlike mammals, avian erythrocytes retain functional mitochondria throughout their lifespan, which is hypothesised to support a higher energetic capacity, thereby enhancing metHb reductase activity. However, the precise metabolic responses of erythrocytes to metHb-inducing agents are not well understood, complicating the development of such toxins. Therefore, this study aimed to explore the real-time effects of metHb-inducing agents on erythrocyte respiration, and to monitor the metHb generation and reduction following agent administration in both avian and mammalian species. Laboratory experiments utilised high-resolution respirometry in conjunction with spectrometry using domestic chickens (Gallus domesticus), laboratory rats (Rattus norvegicus), and a human subject (Homo sapiens, male) as models for avian and mammalian species, respectively, to assess differences in Hb respiration and oxidation states under the influence of metHb-inducing agents. Additionally, field studies employed smartphones to acquire red colour values (RCV) as a potential indicator of metHb levels. Our respirometry results revealed that avian erythrocytes maintained aerobic respiration during Hb oxidation, in contrast to mammalian erythrocytes which rely solely on glycolysis. Moreover, both spectrometry and RCV results showed that avian erythrocytes generated metHb at a significantly slower rate than mammalian erythrocytes under equivalent doses of metHb-inducing agents, demonstrating a higher tolerance. These findings introduce and validate a novel methodology using intact erythrocytes to monitor Hb respiration and oxidation state, providing insights into the distinct metabolic activities in avian and mammalian erythrocytes under oxidative stress, proving their differing capacities to resist such conditions. This methodology is crucial for further identifying synergists or protectants that target specific erythrocyte metabolism to improve the selectivity of current metHb-inducing agents.