Glutamate acts at NMDA receptors on fresh bovine and on cultured human retinal pigment epithelial cells to trigger release of ATP

Efficient vesicle membrane recycling at presynaptic terminals is pivotal for preventing depletion and maintaining high firing rates in neuronal networks. We used a new approach, based on the combination of spectrally different optical probes, to investigate how stimulation determines the fate of synaptic vesicles after endocytosis. We found that in the small central synapses of rat hippocampal neurones low frequency stimulation (40 action potentials at 2 Hz) targets vesicles preferentially to vesicle pools that were kinetically faster. Vesicles taken up during endocytosis triggered by high frequency stimulation (400 action potentials, 20 Hz) were also placed in the back of the release queue. We performed a spatial analysis of the recycled vesicles in living hippocampal boutons using two spectrally different FM-dyes (FM1-43 and FM5-95). By using these consecutively, vesicles endocytosed by either stimulation protocol were labelled with a different colour. This revealed that the kinetic arrangement was also reflected in the spatial organization of vesicles within the bouton. Next, we identified the postsynaptic site of the active zone by transfecting the neurones with postsynaptic density protein PSD-95-CFP. The data from these triple colour experiments suggest that retrieval after low frequency stimulation keeps vesicles in a more confined region closer to the active zone as identified by PSD-95-CFP expression at the postsynaptic site.     

Adding Bupivacaine to High-potassium Cardioplegia Improves Function and Reduces Cellular Damage of Rat Isolated Hearts after Prolonged, Cold Storage

Background: Bupivacaine retards myocardial acidosis during ischemia. The authors measured function of rat isolated hearts after prolonged storage to determine whether bupivacaine improves cardiac protection compared with standard cardioplegia alone.

Methods: After measuring cardiac function on a Langendorff apparatus, hearts were perfused with cardioplegia alone (controls), cardioplegia containing 500 [mu]m bupivacaine, or cardioplegia containing 2 mm lidocaine; were stored at 4[degrees]C for 12 h; and were then reperfused. Heart rate and left ventricular developed pressures were measured for 60 min. Maximum positive rate of change in ventricular pressure, oxygen consumption, and lactate dehydrogenase release were also measured.

Results: All bupivacaine-treated, four of five lidocaine-treated, and no control hearts beat throughout the 60-min recovery period. Mean values of heart rate, left ventricular developed pressure, maximum positive rate of change in ventricular pressure, rate-pressure product, and efficiency in bupivacaine-treated hearts exceeded those of the control group (P < 0.001 at 60 min for all). Mean values of the lidocaine group were intermediate. Oxygen consumption of the control group exceeded the other groups early in recovery, but not at later times. Lactate dehydrogenase release from the bupivacaine group was less than that from the control group (P < 0.001) but did not differ from baseline.     

Nerve growth factor and tyrosine kinase A in human salivary adenoid cystic carcinoma: expression patterns and effects on in vitro invasive behavior.

PURPOSE: Perineural invasion is a frequent occurrence in salivary adenoid cystic carcinoma (ACC) and may prevent complete surgical resection. Studies have indicated that nerve growth factor (NGF) and its high-affinity receptor tyrosine kinase A (TrkA) may play a role in perineural invasion in several malignancies in which perineural invasion is observed. The present study was conducted to investigate the expression of NGF and TrkA in salivary ACC and to examine the effects of NGF on adhesion, migration and invasion capacities of a salivary ACC cell line (SACC-83) in vitro. PATIENTS AND METHODS: Expression of NGF and TrkA was explored using immunohistochemistry in paraffin-embedded tissues of 32 cases of salivary ACC. The effects of NGF on in vitro adhesion, migration, and invasion capacities of the SACC-83 cell line were examined using an MTT assay and a modified Boyden chamber assay respectively. RESULTS: In ACC specimens, 31 (96.9%) and 32 (100%) tumors showed immunoreactivity for NGF and TrkA respectively. Significant correlations were found between NGF/TrkA expression levels and perineural invasion (P < .05). In cell adhesion assay, the percent adherences of SACC-83 cells co-cultured with 25 ng/ml NGF at 1.5 hours and 5, 25 ng/ml NGF at 6 hours were significantly higher than that co-cultured with 0 ng/ml NGF (P < .05). However, high concentration of NGF (500 ng/ml) resulted in a significant inhibition of invasion (P < .05). CONCLUSION: Overexpression of NGF and TrkA in human salivary ACC tissues may constitute a reason for perineural invasion in salivary ACC.     

Role of basal insulin in the regulation of protein kinetics and energy metabolism in septic patients.

We have investigated the role of basal insulin concentration on leucine kinetics (determined by means of 1-[13C]leucine) and energy metabolism (determined by indirect calorimetry) in eight septic patients by reducing insulin (and glucagon) secretion by somatostatin infusion. Basal glucagon concentration was elevated (744 +/- 381 pg/mL), and insulin concentration was normal (10 +/- 4 microU/mL). Basal resting energy expenditure (REE) was 151 +/- 8% that of predicted basal energy expenditure, and leucine appearance (Ra), oxidation, and nonoxidative disposal rates were all elevated above the normal ranges. Somatostatin infusion reduced insulin concentration by 52% and glucagon concentration by 64%. This resulted in a significant increase in the rate of leucine oxidation from 0.96 +/- 0.08 to 1.18 +/- 0.14 mumol/kg/min (p less than 0.01), and nonoxidative leucine disposal decreased from 2.95 +/- 0.18 to 2.67 +/- 0.17 mumol/kg/min (p less than 0.01). Somatostatin infusion also caused significant increases in REE and fat oxidation from 1310 +/- 100 to 1505 +/- 128 kcal/m2/day (p less than 0.05) and from 1.72 +/- 0.24 to 2.41 +/- 0.41 mg/kg/min, respectively, and a slight decrease of carbohydrate oxidation from 1.51 +/- 0.49 to 1.31 +/- 0.49 mg/kg/min. These metabolic responses can be attributed to the reduction in insulin concentration, because they are in the opposite direction of changes that would occur as a consequence of a reduction in glucagon concentration. We conclude that the basal insulin plays an important role in attenuating net protein loss and energy expenditure.     

In Vivo Studies of Pulsatile Implantable Impeller Assist and Total Hearts

Abstract: A pulsatile impeller assist heart and a total heart were tested as a chronic left ventricular assist device in 5 calves and an acute biventricular assist device in 4 pigs respectively, to evaluate their blood compatibility. During the left ventricular assist experiments, the indicators for hemolysis, thrombogenesis, renal dysfunction, and hepatic dysfunction were measured preoperatively, at the beginning of the pumping, 6 h postoperatively, and every following day. The results demonstrated that the impeller assist heart causes no severe blood damage nor organ dysfunction in the experiments lasting up to 11 days. In biventricular assist experiments, the number of red blood cells, white blood cells, platelets, and the he-matocrit, hemoglobin, free hemoglobin, and lactate dehy-drogenase levels were tested preoperatively at the beginning of the pumping and every 2 h postoperatively. The data remained in acceptable ranges during experiments lasting 6 h. It is confirmed that the authors' impeller assist heart and total heart have the advantages of simplicity, implantability, and pulsatility with good blood compatibility.