HIGH-RESOLUTION MICROSCOPIC DETERMINATION OF HEPATIC NADH FLUORESCENCE FOR IN-VIVO MONITORING OF TISSUE OXYGENATION DURING HEMORRHAGIC-SHOCK AND RESUSCITATION

Impaired microvascular oxygen supply reduces oxidative phosphorylation and causes an increase in cellular NADH, which was monitored densitom etrically in vivo by high-resolution fluorescence microscopy (330-390/ >430 nm excitation/emission wavelengths) in rat livers (n = 8) subject ed to hemorrhagic shock and resuscitation. At each time point, NADH fl uorescence was recorded from 10 different observation fields of the le ft liver lobe. Withdrawal of a total of 4.5 mi arterial blood for indu ction of volume-controlled hemorrhagic shade resulted in an increase i n NADH fluorescence by similar to 31% from 45.1 +/- 3.9 to 59.2 +/- 4. 2 aU, which was associated with a all of arterial blood pressure from 110 +/- 3 to 51 +/- 8 mmHg, a decrease in hepatic tissue oxygenation ( flexible polarographic surface electrode) from 18 +/- 2 to 2 +/- I mmH g, and a restriction of hepatic bile flow fi-om 1.7 +/- 0.1 to 0.5 +/- 0.2 mu l/min x g. Normovolemic resuscitation with 10% hydroxyethylsta rch failed to completely restore the metabolic state of liver tissue ( NADH fluorescence 49.9 +/- 3.1 ati), arterial blood pressure (83 +/- 8 mmHg), hepatic tissue oxygenation (7.4 +/- 1.5 mmHg), and hepatocellu lar excretory function (1.3 +/- 0.1 mu l/min x g). During both shock a nd resuscitation, the ratio ber-vc een pericentral and periportal NADH fluorescence intensities slightly increased, but calculation of coeff icients of variance of interlobular NADH fluorescence did not reveal a n increase in heterogeneity of tissue metabolic state. Significant cor relations were found between NADH fluorescence and both hepatic tissue oxygenation (r(2) = 0.78, P < 0.01) and hepatic bile flow (r(2) = 0.8 5, P < 0.01), indicating that high-resolution intravital microscopic a ssessment of NADH fluorescence reflects appropriately the relation bet ween local oxygen supply and demand in hepatic tissue in vivo. (C) 199 7 Academic Press.