LKB1 acts as a critical brake for the glucagon-mediated fasting response.

As important as the fasting response is for survival, an inability to shut it down once nutrients become available can lead to exacerbated disease and severe wasting. The liver is central to transitions between feeding and fasting states, with glucagon being a key initiator of the hepatic fasting response. However, the precise mechanisms controlling fasting are not well defined. One potential mediator of these transitions is liver kinase B1 (LKB1), given its role in nutrient sensing. Here, we show LKB1 knockout mice have a severe wasting and prolonged fasting phenotype despite increased food intake. By applying RNA sequencing and intravital microscopy, we show that loss of LKB1 leads to a dramatic reprogramming of the hepatic lobule through robust up-regulation of periportal genes and functions. This is likely mediated through the opposing effect that LKB1 has on glucagon pathways and gene expression. Conclusion: Our findings show that LKB1 acts as a brake to the glucagon-mediated fasting response, resulting in "periportalization" of the hepatic lobule and whole-body metabolic inefficiency. These findings reveal a mechanism by which hepatic metabolic compartmentalization is regulated by nutrient-sensing.

A Novel Mouse Model to Analyze Non-Genomic ERα Physiological Actions.

Nongenomic effects of estrogen receptor α (ERα) signaling have been described for decades. Several distinct animal models have been generated previously to analyze the nongenomic ERα signaling (eg, membrane-only ER, and ERαC451A). However, the mechanisms and physiological processes resulting solely from nongenomic signaling are still poorly understood. Herein, we describe a novel mouse model for analyzing nongenomic ERα actions named H2NES knock-in (KI). H2NES ERα possesses a nuclear export signal (NES) in the hinge region of ERα protein resulting in exclusive cytoplasmic localization that involves only the nongenomic action but not nuclear genomic actions. We generated H2NESKI mice by homologous recombination method and have characterized the phenotypes. H2NESKI homozygote mice possess almost identical phenotypes with ERα null mice except for the vascular activity on reendothelialization. We conclude that ERα-mediated nongenomic estrogenic signaling alone is insufficient to control most estrogen-mediated endocrine physiological responses; however, there could be some physiological responses that are nongenomic action dominant. H2NESKI mice have been deposited in the repository at Jax (stock no. 032176). These mice should be useful for analyzing nongenomic estrogenic responses and could expand analysis along with other ERα mutant mice lacking membrane-bound ERα. We expect the H2NESKI mouse model to aid our understanding of ERα-mediated nongenomic physiological responses and serve as an in vivo model for evaluating the nongenomic action of various estrogenic agents.

Intravital Microscopy for the Study of Hepatic Glucose Uptake.

The liver is central in maintaining glucose homeostasis. Indeed, impaired hepatic glucose uptake has been implicated in the development of hyperglycemia in type II diabetes (T2D) and non-alcoholic fatty liver disease (NAFLD). However, current approaches to evaluate glucose mobilization rely on indirect measurements that do not provide spatial and temporal information. Here, we describe confocal-based intravital microscopy (IVM) of the liver that allows the identification of hepatocyte spatial organization and glucose transport. Specifically, we describe a method to fluorescently label hepatic landmarks to identify different compartments within the liver. In addition, we outline an in vivo fluorescent glucose uptake assay to quantitatively measure glucose mobilization in space and time. These protocols allow direct investigation of hepatic glycemic control and can be further applied to murine models of liver disease. © Published 2021. This article is a U.S. Government work and is in the public domain in the USA. Basic Protocol 1: Mouse surgical procedure and positioning for liver intravital imaging Basic Protocol 2: Fluorescent labeling and intravital imaging of mouse hepatic compartments Basic Protocol 3: Mouse hepatic glucose uptake assay and intravital imaging analysis.

Experimental models for evaluating non-genomic estrogen signaling.

Non-genomic effects of estrogen receptor α (ERα) signaling have been described for decades. However, the mechanisms and physiological processes resulting solely from non-genomic signaling are poorly understood. Challenges in studying these effects arise from the strongly nucleophilic tendencies of estrogen receptor, and many approaches to excluding ERα from the nucleus have been explored over the years. In this review, we discuss past strategies for studying ERα's non-genomic action and current models, specifically H2NES ERα, first described by Burns et al. (2011). In vitro and preliminary in vivo data from H2NES ERα and H2NES mice suggest a promising avenue for pinpointing specific non-genomic ERα action.