A method for determination in situ of variations within the hepatic lobule of hepatocyte function and metabolite concentrations

Shamus P. Burns, Robert D. Cohen, Richard A. Iles, Jocelyn P. Germain, Thomas C.H. Going, Stephen J.W. Evans, Patrick Royston

Research output: Contribution to journalArticle

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Abstract

A method is described for the production of detailed maps of intralobular variations of hepatocyte function and metabolite concentrations, based on variable destruction by digitonin of the lobule from the centrilobular direction. Instead of the conventional approach, in which isolated hepatocytes are then prepared and studied in suspension, perfusion is continued after digitonin treatment and the function of the unaffected lobular remnants is determined, or mean metabolite concentrations are measured by 31P-NMR. These measurements are plotted against the degree of destruction, determined precisely after each study by automated quantitative histomorphometry. These plots are transformed into curves of the function or metabolite concentration of nominal single cells at any point along the radius of the lobule. Gluconeogenesis from lactate remained stable, although reduced, even after 85-90%, lobular destruction, predominated periportally and disappeared by 50% along the radius of the lobule. In 31P-NMR studies, employing 1.5 mM lactate as substrate, narrowing of the intracellular P1 resonance was observed as digitonin destruction increased; this was attributed to a decrease in the intralobular heterogeneity of the intracellular pH, which fell from approx. 7.9 to < 7.4 along the first 16% of the lobular radius (from the periportal end) and to < 7.3 in the remainder of the lobule. The ATP concentration rose, and then fell, along the radius of the lobule in a centripetal direction. The method is potentially generally applicable to a wide range of hepatocellular functions and to the measurement of metabolite concentrations, most conveniently those susceptible to estimation by NMR.
LanguageEnglish
Pages377-383
Number of pages7
JournalBiochemical Journal
Volume319
Issue number2
DOIs
Publication statusPublished - 15 Oct 1996
Externally publishedYes

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Digitonin
Metabolites
Hepatocytes
Liver
Lactic Acid
Nuclear magnetic resonance
Gluconeogenesis
Suspensions
Perfusion
Adenosine Triphosphate
Substrates
Direction compound

Cite this

Burns, Shamus P. ; Cohen, Robert D. ; Iles, Richard A. ; Germain, Jocelyn P. ; Going, Thomas C.H. ; Evans, Stephen J.W. ; Royston, Patrick. / A method for determination in situ of variations within the hepatic lobule of hepatocyte function and metabolite concentrations. In: Biochemical Journal. 1996 ; Vol. 319, No. 2. pp. 377-383.
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abstract = "A method is described for the production of detailed maps of intralobular variations of hepatocyte function and metabolite concentrations, based on variable destruction by digitonin of the lobule from the centrilobular direction. Instead of the conventional approach, in which isolated hepatocytes are then prepared and studied in suspension, perfusion is continued after digitonin treatment and the function of the unaffected lobular remnants is determined, or mean metabolite concentrations are measured by 31P-NMR. These measurements are plotted against the degree of destruction, determined precisely after each study by automated quantitative histomorphometry. These plots are transformed into curves of the function or metabolite concentration of nominal single cells at any point along the radius of the lobule. Gluconeogenesis from lactate remained stable, although reduced, even after 85-90{\%}, lobular destruction, predominated periportally and disappeared by 50{\%} along the radius of the lobule. In 31P-NMR studies, employing 1.5 mM lactate as substrate, narrowing of the intracellular P1 resonance was observed as digitonin destruction increased; this was attributed to a decrease in the intralobular heterogeneity of the intracellular pH, which fell from approx. 7.9 to < 7.4 along the first 16{\%} of the lobular radius (from the periportal end) and to < 7.3 in the remainder of the lobule. The ATP concentration rose, and then fell, along the radius of the lobule in a centripetal direction. The method is potentially generally applicable to a wide range of hepatocellular functions and to the measurement of metabolite concentrations, most conveniently those susceptible to estimation by NMR.",
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A method for determination in situ of variations within the hepatic lobule of hepatocyte function and metabolite concentrations. / Burns, Shamus P.; Cohen, Robert D.; Iles, Richard A.; Germain, Jocelyn P.; Going, Thomas C.H.; Evans, Stephen J.W.; Royston, Patrick.

In: Biochemical Journal, Vol. 319, No. 2, 15.10.1996, p. 377-383.

Research output: Contribution to journalArticle

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