Cold acclimation preserves hindgut reabsorption capacity at low temperature in a chill-susceptible insect, Locusta migratoria

Highlights

• Cold exposure reduced ion transport across the hindgut of a chill-susceptible insect (L. migratoria).

• Cold acclimation prevented the loss of ion transport.

• cGMP and VHA-dependent ion transport are blocked following cold exposure but preserved by cold acclimation.

• Limited support of homeoviscous adaptation related to changes in hindgut capacity.

Abstract

Cold acclimation increases cold tolerance of chill-susceptible insects and the acclimation response often involves improved organismal ion balance and osmoregulatory function at low temperature. However, the physiological mechanisms underlying plasticity of ion regulatory capacity are largely unresolved. Here we used Ussing chambers to explore the effects of cold exposure on hindgut KCl reabsorption in cold- (11 °C) and warm-acclimated (30 °C) Locusta migratoria. Cooling (from 30 to 10 °C) reduced active reabsorption across recta from warm-acclimated locusts, while recta from cold-acclimated locusts maintained reabsorption at 10 °C. The differences in transport capacity were not linked to major rearrangements of membrane phospholipid profiles. Yet, the stimulatory effect of two signal transduction pathways were altered by temperature and/or acclimation. cAMP-stimulation increased reabsorption in both acclimation groups, with a strong stimulatory effect at 30 °C and a moderate stimulatory effect at 10 °C. cGMP-stimulation also increased reabsorption in both acclimation groups at 30 °C, but their response to cGMP differed at 10 °C. Recta from warm-acclimated locusts, characterised by reduced reabsorption at 10 °C, recovered reabsorption capacity following cGMP-stimulation at 10 °C. In contrast, recta from cold-acclimated locusts, characterised by sustained reabsorption at 10 °C, were unaffected by cGMP-stimulation. Furthermore, cold-exposed recta from warm-acclimated locusts were insensitive to bafilomycin-α1, a V-type H⁺-ATPase inhibitor, whereas this blocker reduced reabsorption across cold-exposed recta from cold-acclimated animals. In conclusion, bafilomycin-sensitive and cGMP-dependent transport mechanism(s) are likely blocked during cold exposure in warm-acclimated animals while preserved in cold-acclimated animals. These may in part explain the large differences in rectal ion transport capacity between acclimation groups at low temperature.

Introduction

Thermal tolerance of insects is of paramount importance for insect species distribution (Kellermann et al., 2012; Sunday et al., 2011) and the effect of low temperatures on insect physiology has therefore received considerable attention (Bale, 1996; Denlinger and Lee, 2010; Sinclair, 1999; Zachariassen, 1985). In many insects, cold tolerance is linked to their ability to preserve ion and water homeostasis at low temperature since “chill-susceptible” insects may experience chill injury/mortality when ion homeostasis is lost (Bayley et al., 2018; Gerber and Overgaard, 2018; Koštál et al., 2004; MacMillan and Sinclair, 2011a; Overgaard and MacMillan, 2017). Accordingly, it is now clear that the capacity of the insect osmoregulatory system to preserve ion and water balance at low temperature is crucial for their cold tolerance and survival (Andersen et al., 2017a; Andersen and Overgaard, 2020; Des Marteaux et al., 2018; Gerber and Overgaard, 2018; Lebenzon et al., 2020; MacMillan et al., 2015a, MacMillan et al., 2015b, MacMillan et al., 2015c; MacMillan and Sinclair, 2011b; Olsson et al., 2016; Yerushalmi et al., 2018).

Cold exposure is associated with a dramatic loss of transport capacity in osmoregulatory organs in locusts (Gerber and Overgaard, 2018) and other orthopterans (Houlihan and Sell, 1984; Williams et al., 1978). Further, chronic cold acclimation or rapid cold hardening improves cold tolerance through increased capacity for ion regulation following cold exposure in L. migratoria (Andersen et al., 2017a; Findsen et al., 2013). The link between cold tolerance and osmoregulatory function in L. migratoria was further strengthened by the finding that cold acclimation increases the capacity of the hindgut to reabsorb fluid and ions at low temperature (Gerber and Overgaard, 2018). However, little is known about the physiological mechanisms underlying the plasticity of osmoregulatory systems at low temperature. There is equivocal evidence that loss of ion and water balance is linked to disruption of passive transport across the gut epithelia of L. migratoria (Brzezinski and MacMillan, 2020; Gerber and Overgaard, 2018). The temperature dependence and sensitivity of active transport processes may also shape the thermal response among acclimation groups. This is supported by data from Des Marteaux et al. (2017), where acclimation induced a shift in gene transcription of rectal ion transporters [Na⁺/K⁺ ATPase (NKA) and V-type H⁺-ATPase (VHA)], and genes involved in cAMP and cGMP signal transduction pathways in the hindgut of the fall field cricket. Our previous physiological study of hindgut reabsorption in L. migratoria also demonstrated that the loss of osmoregulatory capacity observed across the hindgut of warm-acclimated locusts was primarily caused by reduced active transport (Gerber and Overgaard, 2018). It is possible that temperature effects on membrane fluidity compromise the function of transmembrane ion transporters and hence the rate of passive and/or active ion transport (Hazel, 1989, Hazel, 1995). Cold acclimation (which enhances cold tolerance) is often associated with cell membrane restructuring in insects (Colinet et al., 2016; Enriquez and Colinet, 2019; Koštál, 2010), but modifications in membrane lipid composition during cold acclimation remain largely unexplored in Orthopterans (but see Bayley et al., 2020).

To identify the physiological mechanism(s) involved in rectal transport during cold exposure and cold acclimation, the present study explored the regulation of KCl reabsorption across the rectum at high (30 °C) and low (10 °C) temperature in warm- and cold-acclimated L. migratoria, known to differ in their cold tolerance (Andersen et al., 2017a). Using Ussing chambers, we investigated 1) if there was a loss of active transport across the rectum of the chill-susceptible migratory locust at low temperature; 2) whether cold acclimation aided to preserve active transport; 3) if thermal sensitivity of cAMP and cGMP stimulation of rectal KCl transport was involved in the acclimation response and 4) whether thermal acclimation changed the contribution of VHA in active transport at low temperature. Finally, we compared the phospholipid composition of the hindgut from cold- and warm-acclimated locusts to; 5) determine whether cold acclimation induces membrane remodelling (i.e., homeoviscous adaptation). With these experiments we were able to identify a number of physiological mechanisms responsible for the apparent differences in ionoregulatory capacity that exist between cold- and warm-acclimated locusts exposed to low temperatures.