Dynamics as well as localization of Ca2+ transients plays a vital role in liver function under homeostatic conditions, repair, and disease. In response to circulating hormonal stimuli, hepatocytes exhibit intracellular Ca2+ responses that propagate through liver lobules in a wave-like fashion. Although intracellular processes that control cell autonomous Ca2+ spiking behavior have been studied extensively, the intra- and inter-cellular signaling factors that regulate lobular scale spatial patterns and wave-like propagation of Ca2+ remain to be determined. To address this need, we acquired images of cytosolic Ca2+ transients in 1300 hepatocytes situated across several mouse liver lobules over a period of 1600 s. We analyzed this time series data using correlation network analysis, causal network analysis, and computational modeling, to characterize the spatial distribution of heterogeneity in intracellular Ca2+ signaling components as well as intercellular interactions that control lobular scale Ca2+ waves. Our causal network analysis revealed that hepatocytes are causally linked to multiple other co-localized hepatocytes, but these influences are not necessarily aligned uni-directionally along the sinusoids. Our computational model-based analysis showed that spatial gradients of intracellular Ca2+ signaling components as well as intercellular molecular exchange are required for lobular scale propagation of Ca2+ waves. Additionally, our analysis suggested that causal influences of hepatocytes on Ca2+ responses of multiple neighbors lead to robustness of Ca2+ wave propagation through liver lobules.

Objective The purpose of the present study is to model the dynamics of lobular Ca2+ wave propagation induced by an extracellular stimulus and to analyze the effect of spatially systematic variations in cell-intrinsic signaling parameters on sinusoidal Ca2+ response. Methods We developed a computational model of lobular scale Ca2+ signaling that accounts for receptor-mediated initiation of cell-intrinsic Ca2+ signal in hepatocytes and its propagation to neighboring hepatocytes through gap junction-mediated molecular exchange. Results Analysis of the simulations showed that a pericentral-to-periportal spatial gradient in hormone sensitivity and/or rates of IP3 synthesis underlies the Ca2+ wave propagation. We simulated specific cases corresponding to localized disruptions in the graded pattern of these parameters along a hepatic sinusoid. Simulations incorporating locally altered parameters exhibited Ca2+ waves that do not propagate throughout the hepatic plate. Increased gap junction coupling restored normal Ca2+ wave propagation when hepatocytes with low Ca2+ signaling ability were localized in the mid-lobular or the pericentral region. Conclusion Multiple spatial patterns in intracellular signaling parameters can lead to Ca2+ wave propagation that is consistent with the experimentally observed spatial patterns of Ca2+ dynamics. Based on simulations and analysis, we predict that increased gap junction-mediated intercellular coupling can induce robust Ca2+ signals in otherwise poorly responsive hepatocytes, at least partly restoring the sinusoidally oriented Ca2+ waves. Significance Our bottom-up model of agonist-evoked spatial Ca2+ patterns can be integrated with detailed descriptions of liver histology to study Ca2+ regulation at the tissue level.

The analysis of molecular states of individual cells, as defined by their mRNA expression profiles and protein composition, has gained widespread interest in studying biological phenomena ranging from embryonic development to homeostatic tissue function and genesis and evolution of cancers. Although the molecular content of individual cells in a tissue can vary widely, their molecular states tend to be constrained within a transcriptional landscape partly described by the canonical archetypes of a population of cells. In this study we sought to characterize the effects of an acute (partial hepatectomy) and chronic (alcohol consumption) perturbation on the molecular states of individual hepatocytes during the onset and progression of liver regeneration. We analyzed the expression of 84 genes across 233 individual hepatocytes acquired using laser capture microdissection. Analysis of this single cell data revealed that hepatocyte molecular states can be considered as distributed across a set of four states irrespective of perturbation, with the proportions of hepatocytes in these states being dependent on the perturbation. In addition to the quiescent, primed, and replicating hepatocytes, we identified a fourth molecular state lying between the primed and replicating subpopulations. Comparison of the proportions of hepatocytes from each experimental condition in these four molecular states suggested that, in addition to aberrant priming, a slower transition from primed to replication state could contribute towards ethanol-mediated suppression of liver regenerative response to partial hepatectomy.

Rapid breakdown of hepatic glycogen stores into glucose plays an important role during intense physical exercise to maintain systemic euglycemia. Hepatic glycogenolysis is governed by several different liver-intrinsic and systemic factors such as hepatic zonation, circulating catecholamines, hepatocellular calcium signaling, hepatic neuroanatomy, and the central nervous system (CNS). Of the factors regulating hepatic glycogenolysis, the extent of lobular innervation varies significantly between humans and rodents. While rodents display very few autonomic nerve terminals in the liver, nearly every hepatic layer in the human liver receives neural input. In the present study, we developed a multi-scale, multi-organ model of hepatic metabolism incorporating liver zonation, lobular scale calcium signaling, hepatic innervation, and direct and peripheral organ-mediated communication between the liver and the CNS. We evaluated the effect of each of these governing factors on the total hepatic glucose output and zonal glycogenolytic patterns within liver lobules during simulated physical exercise. Our simulations revealed that direct neuronal stimulation of the liver and an increase in circulating catecholamines increases hepatic glucose output mediated by mobilization of intracellular calcium stores and lobular scale calcium waves. Comparing simulated glycogenolysis between human-like and rodent-like hepatic innervation patterns (extensive vs. minimal) suggested that propagation of calcium transients across liver lobules acts as a compensatory mechanism to improve hepatic glucose output in sparsely innervated livers. Interestingly, our simulations suggested that catecholamine-driven glycogenolysis is reduced under portal hypertension. However, increased innervation coupled with strong intercellular communication can improve the total hepatic glucose output under portal hypertension. In summary, our modeling and simulation study reveals a complex interplay of intercellular and multi-organ interactions that can lead to differing calcium dynamics and spatial distributions of glycogenolysis at the lobular scale in the liver.