Glucose transporter expression and regulation following a fast in the ruby-throated hummingbird, Archilochus colubris

Abstract

Hummingbirds, subsisting almost exclusively on nectar sugar, face extreme challenges to blood sugar regulation. The capacity for transmembrane sugar transport is mediated by the activity of facilitative glucose transporters (GLUTs) and their localisation to the plasma membrane (PM). In this study, we determined the relative protein abundance of GLUT1, GLUT2, GLUT3 and GLUT5 via immunoblot using custom-designed antibodies in whole-tissue homogenates and PM fractions of flight muscle, heart and liver of ruby-throated hummingbirds (Archilochus colubris). The GLUTs examined were detected in nearly all tissues tested. Hepatic GLUT1 was minimally present in whole-tissue homogenates and absent win PM fractions. GLUT5 was expressed in flight muscles at levels comparable to those of the liver, consistent with the hypothesised uniquely high fructose uptake and oxidation capacity of hummingbird flight muscles. To assess GLUT regulation, we fed ruby-throated hummingbirds 1 mol l^-1 sucrose ad libitum for 24 h followed by either 1 h of fasting or continued feeding until sampling. We measured relative GLUT abundance and concentration of circulating sugars. Blood fructose concentration in fasted hummingbirds declined (∼5 mmol l^-1 to ∼0.18 mmol l^-1), while fructose-transporting GLUT2 and GLUT5 abundance did not change in PM fractions. Blood glucose concentrations remained elevated in fed and fasted hummingbirds (∼30 mmol l^-1), while glucose-transporting GLUT1 and GLUT3 in flight muscle and liver PM fractions, respectively, declined in fasted birds. Our results suggest that glucose uptake capacity is dynamically reduced in response to fasting, allowing for maintenance of elevated blood glucose levels, while fructose uptake capacity remains constitutively elevated promoting depletion of blood total fructose within the first hour of a fast.

Keywords: Flight muscle; Fructose; GLUT; Liver; Plasma membrane.

Fig. 1.

Mean concentration of circulating sugars and metabolites. (A) Glucose and (B) fructose concentrations from plasma samples and (C) lactate and (D) pyruvate concentrations from flight muscle homogenates of fed (n=6) and fasted (n=5) male hummingbirds. Data are presented as means±s.e.m. (*P=0.001).

Fig. 2.

Relative protein abundance of GLUT1 in hummingbird flight muscle, heart and liver tissue. Data represent mean±s.e.m. arbitrary units (a.u.) of intensity based on analyses of normalised immunoblots. Ad libitum fed and 1 h fasted hummingbird GLUT1 abundance was measured in (A) whole-tissue homogenates and (B) plasma membrane (PM) fraction samples. Asterisks indicate a significant difference (P<0.05) in GLUT1 between fed and fasted conditions within that tissue (summarised in Tables 1 and 2; Figs S1 and S2). Letters (a,b,c) over individual bars represent a significant difference (P<0.05) of GLUT1 abundance between the specified tissue under the treatment condition (summarised in Tables 3 and 4). Sample sizes are given in the bars for each tissue and treatment.

Fig. 3.

Relative protein abundance of GLUT2 in hummingbird flight muscle, heart and liver tissue. Data represent mean±s.e.m. arbitrary units (a.u.) of intensity based on analyses of normalised immunoblots. Ad libitum fed and 1 h fasted hummingbird GLUT2 abundance was measured in (A) whole-tissue homogenates and (B) PM fraction samples. Asterisks indicate a significant difference (P<0.05) in GLUT2 between fed and fasted conditions within that tissue (summarised in Tables 1 and 2; Figs S1 and S2). Differences in GLUT2 abundance between tissues within the treatment condition are summarised in Tables 3 and 4). Sample sizes are given in the bars for each tissue and treatment.

Fig. 4.

Relative protein abundance of GLUT3 in hummingbird flight muscle, heart and liver tissue. Data represent mean±s.e.m. arbitrary units (a.u.) of intensity based on analyses of normalised immunoblots. Ad libitum fed and 1 h fasted hummingbird GLUT3 abundance was measured in (A) whole-tissue homogenates and (B) PM fraction samples. Asterisks indicate a significant difference (P<0.05) in GLUT3 between fed and fasted conditions within that tissue (summarised in Tables 1 and 2; Figs S1 and S2). Letters (a,b) over individual bars represent a significant difference (P<0.05) of GLUT3 abundance between the specified tissue under the treatment condition (summarised in Tables 3 and 4). Sample sizes are given in the bars for each tissue and treatment.

Fig. 5.

Relative protein abundance of GLUT5 in hummingbird flight muscle, heart and liver tissue. Data represent mean±s.e.m. arbitrary units (a.u.) of intensity based on analyses of normalised immunoblots. Ad libitum fed and 1 h fasted hummingbird GLUT5 abundance was measured in (A) whole-tissue homogenates and (B) PM fraction samples. Differences in GLUT5 abundance between fed and fasted conditions within a given tissue are summarised in Tables 1 and 2 (see also Figs S1 and S2). Differences in GLUT5 abundance between tissues within the treatment condition are summarised in Tables 3 and 4. Sample sizes are given in the bars for each tissue and treatment.