Restricted feeding improves postischemic recovery of Langendorff-perfused rat hearts.

The goal of the present study was to assess the effects of a restricted feeding schedule (RFS) on postischemic contractile recovery in relation to triacylglycerol (TAG), glycogen, and ATP content. Glucose-6-phosphate dehydrogenase (G6PDH) activity, reduced/oxidized glutathione ratio (GSH/GSSG), and thiobarbituric acid reactive substances (TBARS) levels were also determined. Isolated rat hearts entrained to daily RFS (2 h food access starting at 1200) or fed ad libitum (FED) for 3 weeks were Langendorff-perfused (25 min ischemia, 30 min reperfusion) with Krebs-Ringer bicarbonate solution (10 mmol/L glucose). RFS improved the recovery of contractility and reduced creatine kinase (CK) release upon reperfusion. Further, at the end of reperfusion, RFS hearts exhibited increased G6PDH activity and repletion of tissue glycogen, TAG, and ATP that was not observed in the FED hearts. GSH/GSSG at the end of reperfusion fell to the same value in both nutritional states, and TBARS levels were higher in the RFS hearts. In conclusion, RFS improved postischemic functional recovery, which was accompanied by a reduction in CK release and a striking energy recovery. Although enhanced G6PDH activity was displayed, RFS was unable to reduce lipid peroxidation, supporting a clear dissociation between protection against mechanical dysfunction and CK release on the one hand and oxidative damage on the other.

Involvement of mitochondrial permeability transition, glutathione status, pentose phosphate pathway and oxidative damage in the protective effect of fasting on the ischaemic-reperfused rat heart.

1. Fasting, which increases the catabolism of fatty acids, gives functional protection to the ischaemic-reperfused heart. To obtain further knowledge of this cardioprotective effect, changes in mitochondrial permeability transition (MPT) were measured by the entrapment of 2-deoxy-[(3)H]-glucose (2-DG). We also assessed whether MPT is associated with changes in glutathione status, the activity of glucose-6-phosphate-dehydrogenase (G6PDH) and tissue oxidative damage, estimated by the measurement of Thiobarbituric acid-reactive substances (TBARS). 2. Spontaneously beating hearts of fed and 24 h fasted rats were Langendorff perfused with Krebs'-Ringer bicarbonate solution (10 mmol/L glucose) and exposed to 25 min global ischaemia, followed by 30 min reperfusion. 3. Ischaemia-reperfusion resulted in a fourfold increase in mitochondrial entrapment of 2-DG in the fed group. This response was 29% lower in the fasted group, but there were no concomitant changes in total retention of 2-DG in the heart. Fasting increased the activity of G6PDH by a factor of 1.4 and caused a 2.8-fold increase in the ratio of reduced glutathione to oxidized glutathione (GSH:GSSG) at the end of the pre-ischaemic period. Ischaemia-reperfusion did not affect G6PDH activity, but reduced the GSH:GSSG ratio in both the fed and fasted groups by 50%. Therefore, the GSH:GSSG ratio remained higher in the fasted group. Fasting also decreased cellular levels of TBARS by approximately 25%. Lipolysis of endogenous triacylglycerol was increased during the pre-ischaemic period in the fasted group. 4. These data suggest that the enhancement of fatty acid catabolism that occurs in fasting activates mechanisms that tend to reduce oxidative damage and limit MPT.

Protection of ischaemic-reperfused rat heart by dimethylamiloride is associated with inhibition of mitochondrial permeability transition.

1. The aim of the present study was to assess whether protection afforded by the Na(+)/H(+) exchanger blocker dimethylamiloride (DMA) is associated with inhibition of mitochondrial permeability transition (MPT). The effects of DMA were compared with those of cyclosporine (Cs) A, an inhibitor of MPT. 2. Rat hearts were Langendorff perfused with Krebs'-bicarbonate medium containing 10 mmol/L glucose and were subjected to 25 min no-flow global ischaemia and 30 min reperfusion in the presence or absence of 10 micromol/L DMA or 0.2 micromol/L CsA. Cell viability was measured using tetrazolium stain. The MPT was determined by loading hearts with 2-deoxy-[(3)H]-glucose (2DG), which enters mitochondria only during MPT. Total heart 2DG content as an estimation of the extent of tissue damage was also measured. To assess whether DMA has any direct effect on glycolysis, a cell-free heart extract containing all the glycolytic enzymes was used. 3. Dimethylamiloride improved functional recovery (rate-pressure product) from 24 +/- 7 to 68 +/- 11% (P < 0.01) at reperfusion end, attenuated the increase in left ventricular end-diastolic pressure (from 29 +/- 7 to 6 +/- 3% 10 min after reperfusion onset; P < 0.01), improved cell viability (from 21.2 +/- 6.6 to 69.6 +/- 7.1% at reperfusion end; P < 0.05) and lessened lactate accumulation at the end of ischaemia (119 +/- 15 vs 163 +/- 14 micromol/g dry weight; P < 0.05). Dimethylamiloride limited MPT: 2DG mitochondrial entrapment, being 33.1 +/- 14.2 and 96.3 +/- 14.0 at reperfusion end in the treated and control hearts, respectively (P < 0.05), and concomitantly raised total 2DG content (51.3 +/- 4.4 vs 86.8 +/- 1.7 x 10(3) d.p.m./g wet weight in control and treated groups, respectively; P < 0.05). Cyclosporine A improved functional recovery and attenuated the amplitude of ventricular diastolic pressure in ischaemic-reperfused hearts. It also reduced mitochondrial entrapment (67.3 +/- 7.7%; P < 0.05 vs control) and increased total cell 2DG content (162.3 +/- 1.3 x 10(3) d.p.m./g wet weight; P < 0.01 vs control) at the end of reperfusion. Dimethylamiloride did not affect glucose consumption and lactate production in the cell-free heart extract. 4. In conclusion, DMA protects against the noxious effects of ischaemia-reperfusion and inhibits MPT, coinciding with present and previous findings concerning the effects of CsA. Dimethylamiloride also diminished lactate accumulation, although it did not exhibit any direct effect on glycolysis. These data suggest that blockade of Na(+)/H(+) exchange by DMA attenuates the extent of MPT in ischaemic-reperfused rat heart.

Role of endogenous nitric oxide in classic preconditioning in rat hearts.

Ischemic preconditioning (IPC) protects the heart against subsequent sustained ischemia reperfusion (RP). Despite many triggers and signaling pathways, which seem to be involved in IPC, the IPC-mechanisms remain a controversial issue. One of them is endogenous production of nitric oxide (NO). To assess the role of NO in IPC and its relation with glycogen and glycolysis, the effects of inhibiting NO synthase with L-NAME (50 microM) were examined in IPC rat hearts perfused with medium containing 10 mM glucose. Left ventricular developed pressure-rate product (RPP) and end diastolic pressure (EDP), lactate and glycogen contents, and cell viability were measured. Global ischemia (25 min) was followed by 30 min RP. IPC consisted in one cycle of 3 min ischemia-5 min RP. IPC reduced EDP and improved RP recovery of RPP. L-NAME had no effects on the non-IPC group but abolished these effects of IPC. IPC reduced ischemic decrease of glycogen and the acceleration of glycolysis, and improved cell viability. L-NAME did not affect these effects of IPC. The results suggest that NO is ineffective on the noxious effects of ischemia-RP in non-IPC hearts and on the effects of IPC on cell viability, glycogenolysis and glycolysis whereas it is only involved in functional protection.