Thermal acclimation mitigates cold-induced paracellular leak from the Drosophila gut

Abstract

Chill susceptible insects suffer tissue damage and die at low temperatures. The mechanisms that cause chilling injury are not well understood but a growing body of evidence suggests that a cold-induced loss of ion and water homeostasis leads to hemolymph hyperkalemia that depolarizes cells, leading to cell death. The apparent root of this cascade is the net leak of osmolytes down their concentration gradients in the cold. Many insects, however, are capable of adjusting their thermal physiology, and cold-acclimated Drosophila can maintain homeostasis and avoid injury better than warm-acclimated flies. Here, we test whether chilling causes a loss of epithelial barrier function in female adult Drosophila, and provide the first evidence of cold-induced epithelial barrier failure in an invertebrate. Flies had increased rates of paracellular leak through the gut epithelia at 0 °C, but cold acclimation reduced paracellular permeability and improved cold tolerance. Improved barrier function was associated with changes in the abundance of select septate junction proteins and the appearance of a tortuous ultrastructure in subapical intercellular regions of contact between adjacent midgut epithelial cells. Thus, cold causes paracellular leak in a chill susceptible insect and cold acclimation can mitigate this effect through changes in the composition and structure of transepithelial barriers.

Chilling induces unidirectional solute leak through the locust gut epithelia

“…Similar to trends observed in Drosophila , we predicted that a large and rapid increase in gut FITC-dextran concentration (i.e. serosal to mucosal leak) would occur over time in the cold (MacMillan et al, 2017). However, when each gut segment was analysed for marker content, we found that cold stress induced minimal leak from the hemolymph into the gut (Fig.…”

“…To measure paracellular permeability in the gut epithelia of locusts, we monitored the movement of a fluorescently-labeled molecule in both the serosal (hemolymph) to mucosal (lumen) direction, and the mucosal to serosal direction. All experiments used FITC-dextran (3-5kDa, Sigma Aldrich, St. Louis, USA) a commonly used probe for determining paracellular permeability in both invertebrate and vertebrate models such as fruit flies ( D. melanogaster ), rats ( Rattus norvegicus domesticus ), and zebra fish ( Danio rerio ) (see MacMillan et al, 2017; Condette et al, 2014; Bagnat et al, 2007).…”

“…Notably, flies acclimated to colder conditions are more cold tolerant than warm-acclimated flies and upregulate approximately 60% of genes encoding known or putative fly SJ proteins (MacMillan et al, 2016b). These cold-acclimated (10°C) flies also have reduced rates of paracellular leak of a fluorescent probe (FITC-dextran) from their gut lumen to their hemolymph, both before and during a cold stress, compared to warm-acclimated (25°C) flies (MacMillan et al, 2017). Together, these studies suggest that cold exposure can cause increased rates of leak through the paracellular barriers, and that improvements in cold tolerance may be in-part related to an improved ability to maintain paracellular barriers.…”

“…The former are typically observed in ectodermally-derived epithelia such as the foregut and hindgut, and are 2-3 nm wide, while smooth SJs are found in endodermally-derived tissues like the midgut, and are 5-20 nm wide (Jonusaite et al, 2016). To date, the majority of SJ studies has been conducted on Drosophila , including the identification of SJ types, associated proteins, and SJ influence in cold tolerance plasticity (Izumi and Furuse, 2014; Jonusaite et al, 2016; MacMillan et al, 2017). Notably, flies acclimated to colder conditions are more cold tolerant than warm-acclimated flies and upregulate approximately 60% of genes encoding known or putative fly SJ proteins (MacMillan et al, 2016b).…”

Cold stress results in sustained locomotor and behavioral deficits in Drosophila melanogaster

“…Jonusaite, Kelly, & Donini, 2017). In D. melanogaster male reproductive behaviors are negatively influenced by severe cold exposure (Singh & Prasad, 2016), but exposure to brief cold acclimation (i.e., rapid cold hardening) preserves male reproductive behavior and success (Shreve et al, 2004).…”