Ezio Vincenti

Influence of the anesthetic 2,6-diisopropylphenol on the oxidative phosphorylation of isolated rat liver mitochondria.

Isolated rat liver mitochondria have been incubated in the presence of the general anesthetic 2,6-diisopropylphenol (0-100 microM) and the efficiency of oxidative phosphorylation has been evaluated by measuring the respiratory rates, the rates of ATP synthesis or hydrolysis and the magnitude of the transmembrane electrical potential. The results obtained indicate that: (a) in mitochondria energized either by succinate or by ATP, 2,6-diisopropylphenol decreased the transmembrane electrical potential and increased the rates of either electron transfer or ATP hydrolysis; (b) in succinate-energized mitochondria 2,6-diisopropylphenol, at concentrations causing substantial depression of the transmembrane electrical potential, did not modify either the rate of phosphorylation of added ADP or the rate of ADP-stimulated respiration: (c) in succinate-energized mitochondria 2,6-diisopropylphenol caused a concentration-dependent inhibition of the uncoupler-stimulated rate of succinate oxidation. These findings suggest that under the experimental conditions reported 2,6-diisopropylphenol affected the generation and/or maintenance of the transmembrane electrical potential while leaving unchanged the coupling between the electron flow in the respiratory chain and the synthesis of ATP.

Uncoupling effect of the general anesthetic 2,6-diisopropylphenol in isolated rat liver mitochondria

2,6-Diisopropylphenol, a general anesthetic, was previously reported to reduce the transmembrane electrical potential in isolated rat liver mitochondria without affecting the rate of ATP production. This effect appeared to contrast with the generally accepted chemiosmotic mechanism for oxidative phosphorylation. In this study we further examined the influence of 2,6-diisopropylphenol on the production of ATP by isolated mitochondria and we studied its effect on the permeability of the inner mitochondrial membrane to protons. In order to clarify the effects of 2,6-diisopropylphenol on mitochondrial ATP production the activities of the adenine nucleotide translocator and the ATP synthetase were evaluated. The results obtained indicate that the depression of the transmembrane electrical potential elicited by 2,6-diisopropylphenol decreased the activity of the ATP synthetase (as expected in the chemiosmotic model for energy coupling), but not that of the adenine nucleotide translocator. The decrease of the ATP synthetase activity, however, did not result in an apparent inhibition of the overall rate of ATP production in isolated mitochondria due to the rate-limiting effect of the adenine nucleotide translocator in this process. Moreover 2,6-diisopropylphenol was found to increase the permeability to protons of the inner mitochondrial membrane; this effect became more marked as the pH of the incubation medium was increased, demonstrating that it involved the dissociated form of 2,6-diisopropylphenol. These observations suggested that 2,6-diisopropylphenol affected oxidative phosphorylation by acting as a mild protonophore and that its effectiveness was limited by the low fraction of phenol dissociated at near-physiological pH.

Influence of the anesthetic 2,6-diisopropylphenol (propofol) on isolated rat heart mitochondria

The influence of the anesthetic 2,6-diisopropylphenol on isolated rat heart mitochondria has been investigated at a range of concentrations encompassing high and low clinical values. Low clinical concentrations of the anesthetic appeared unable to affect both oxidative phosphorylation and calcium homeostasis. 2,6-diisopropylphenol at high clinical levels decreased both the transmembrane electrical potential and the synthesis of ATP, while leaving mitochondrial calcium homeostasis unaffected. The results obtained suggest that isolated heart mitochondria are substantially insensitive to low clinical concentrations of 2,6-diisopropylphenol, thus largely excluding the possibility that mitochondrial alterations might be involved in the cardiac depression induced by this anesthetic.

Effects of nitrosopropofol on mitochondrial energy-converting system.

Nitrosopropofol (NOPR) is a relatively stable compound obtained from the reaction between the general anesthetic 2,6 diisopropylphenol (propofol) and nitrosoglutathione (GSNO) and bearing a more acidic phenol group than propofol. It interfered with mitochondrial energetic metabolism in a concentration-dependent manner. Concentrations as high as 100 or 200 microM disrupted both oxidative phosphorylation and electron transport. Low concentrations of NOPR (50 microM) markedly slowed down the electron transport rate which was insensitive both to ADP and uncoupler stimulation and spontaneously gradually stopped. Consequently, both the transmembrane potential production and the ATP synthesis system were affected. In the presence of 10 or 20 microM NOPR, mitochondria respired but showed a worsening of the respiratory control and produced a transmembrane potential useful to respond to a phosphorylation pulse, but were not able to restore it. These results were consistent with ATP synthesis and swelling experiments. NOPR was effective at concentrations lower than those required by the combination of propofol and GSNO, suggesting that mitochondria might be able to catalyze the reaction between GSNO and propofol and that the resulting metabolite was more active on mitochondrial membrane structure than the parent compounds. Although the details of the process are yet unknown, the mechanism presented may be of potential relevance to rationalize the pathophysiological effects of propofol.

Differences of ropivacaine and bupivacaine relevant to antiinflammatory actvity, platelet aggregation and antioxidant activity in vitro

Ropivacaine and bupivacaine affect the in vitro growth of rat fibroblasts and monkey kidney Vero cells with bupivacaine generally showing the stronger effect. Up to 3 mM concentration the two anesthetics affect the expression of genes differently for CD2, CD3 gamma, CD40L, IL-2, IL-2R alpha, IL-2R beta, IL-2R gamma, IL-4, and IL-4R during activation of human lymphocytes, with bupivacaine showing the higher effect. Human platelet aggregation is inhibited by the two anesthetics which also show an antioxidant effect on lipid peroxidation of rat liver microsomes. In both cases bupivacaine seems more active than ropivacaine.

Effect of the intravenous anesthetic 2,6-diisopropylphenol on respiration and energy production by rat brain synaptosomes.

The sensitivity of the mitochondrial energy production system to propofol (DPP) has been investigated in rat brain synaptosomes. DPP at 0.8 mM concentration produced a partial inhibition of coupled respiration, an apparent decrease of the oxygen uptake stimulation induced by CCCP and a full inhibition of the mitochondrial ATP production by synaptosomes. Higher concentrations of DPP (1 mM) fully abolish uncoupler-dependent stimulation and at 1.3 mM DPP also coupled respiration is completely blocked. Similar results were obtained when dinitrophenol replaced CCCP and phenol or propylbenzene replaced DPP. The presence of the alkyl residues seems critical for the DPP effect. In the presence of 30 mM glutamate both respiration and ATP production are enhanced but DPP effects are similar to those obtained in the absence of glutamate.

Involvement of long-chain acyl CoA in the antagonistic effects of halothane and L-carnitine on mitochondrial energy-linked processes

Incubation of rat liver mitochondria in the presence of halothane induced a consistent impairment of mitochondrial oxidative phosphorylation without significantly affecting the steady-state of transmembrane electrical potential. These alterations of mitochondrial energy-linked processes were associated with a consistent accumulation of long-chain acyl CoA. Addition of L-carnitine partially prevented the effects of halothane on oxidative phosphorylation and completely abolished the halothane-induced long-chain acyl CoA accumulation. The possibility is discussed that the damaging action of halothane on mitochondrial functions might be partially ascribed to the noxious action of the excess of long-chain acyl CoA induced the anesthetic.

L-carnitine effect on halothane-treated mitochondria.

Addition of halothane to the incubation medium is shown to lower respiratory control and transmembrane potential and to increase ATPase activity in isolated rat liver mitochondria. Evidence is presented that L-carnitine is able to substantially decrease the negative effects of halothane on the energy-linked processes of mitochondria. The effects of halothane and the protective action of L-carnitine are discussed in the light of a possible involvement of long-chain acyl CoA in the unpairing of mitochondrial energy-linked functions.

Mitochondrial effects of L-ropivacaine, a new local anesthetic.

The effects of the local anesthetics ropivacaine and bupivacaine were investigated on isolated rat liver mitochondria. The efficiency of oxidative phosphorylation was evaluated by measuring the rates of respiration and ATP synthesis and the magnitude of the transmembrane electrical potential (deltapsi). Bupivacaine did not alter the ADP-stimulated respiration but strongly affected the resting respiration, which was more than doubled at 0.6 mM. In addition, it decreased the transmembrane electrical potential, and the ATP synthesis rate (deltapsi was less than 100 mV at 0.6 mM). Ropivacaine did not alter the ADP-stimulated respiration, and the resting respiration seemed to be substantially unaffected up to 1.2 mM; a slight increase was observed at 1.8 and 2.4 mM. The transmembrane potential was decreased by anesthetic concentrations higher than 1.2 mM and ATP synthesis was consequently affected. The findings suggest that ropivacaine is less toxic than bupivacaine, in rat liver mitochondria.

Comparison study between acupuncture and pentazocine analgesic and respiratory post-operative effects.

The authors studied the effects of acupuncture and pentazocine on post-operative respiratory function and pain management on patients hysterectomized by means of a subumbilical midline incision. The acupunctural technique consisted of GB-26, St-36, Sp-6 and auricular Shen-Men bilateral electrostimulation for 40 minutes. The analgesic effect of acupuncture was equivalent to that of 30 mg pentazocine, yet the most important effect of acupuncture consisted in a net increase of vital capacity during the period of acupuncture analgesia that lasted for 3-4 hr after stimulation; contrariwise, pentazocine did not cause any vital capacity increment and vital capacity remained at the levels observed prior to narcotic administration.