In the course of our other recent studies, we found 4 such animals. Following sacrifice and subsequent tissue collection, we noted a seemingly abnormal appearance to their bile (Fig. 1A). While other winter squirrels (torpid – T and interbout-aroused – IBA) consistently had deep green bile, the squirrels that failed to hibernate (deemed abnormal – AB) had bright yellow, almost fluorescent bile despite having been sampled at the same time of year. These squirrels had little to any gut contents consistent with the anorexia normally associated with the hibernation season. As indicated in Fig. 1A, collected bile volumes were quite varied

throughout the year but rarely exceeded 500 μl. However, approximately 2.5 ml of bile was collected for one AB animal (Fig. 1A- right). Summer active (SA) squirrels had more varied bile colors as might be expected GSK872 given the effect of diet on bile color [11, 12]. However, our sampling of squirrels from early spring to late summer revealed no simple association of bile color with a given time period (Fig. 1A). Spectral analyses revealed that bile from T and IBA animals contained a peak at approximately 350–500 nm that was not present in either SA or AB squirrels (Fig. 1B). Remarkably, despite having Osimertinib chemical structure a seemingly fluorescent yellow outward appearance,

AB bile was relatively inactive spectrophotometrically. The Exoribonuclease wide range of bile color in summer had little effect on spectral characteristics (data not shown). Figure 1 Gallbladder bile color varies by season and hibernation. A) Photograph of bile collected from golden-mantled ground squirrels (Spermophilus lateralis) as a function of state. Bile was collected from squirrels collected monthly (2–3 squirrels per month) from May (left) until September (right; summer active, SA), squirrels during winter that were torpid (T) when body temperature was ~5°C, squirrels during the euthermic period between bouts

of torpor (interbout-aroused; IBA), and squirrels that were sampled in winter but had failed to hibernate (abnormal, AB). As an indication of approximate volumes, microcentrifuge tubes S63845 contain all of the collected bile for each animal except one AB animal (full tube on lower right; ~2.5 ml of bile was collected from that animal). B) Spectral characteristics of bile as a function of state. Each line represents one animal. Data are depicted for 3 animals of each state and only every 50th symbol is plotted for clarity. Bile acids are produced in the liver by the oxidation of cholesterol and serve important roles in eliminating cholesterol from the body and the emulsification of lipids [13, 14]. Under normal physiological conditions, most bile acids are reabsorbed from the ileum and therefore values typically represent the reabsorption kinetics of bile acids as a function of enterohepatic circulation.