Introduction of macromolecules into synaptosomes using electroporation.

Synaptic terminals are sites of high metabolic activity and thus are particularly vulnerable to oxidative stress. Oxidative damage to proteins can be toxic to neurons and may cause irreversible cell damage and neurodegeneration. A neuroprotective mechanism used by cells to combat oxidative damage is to selectively degrade damaged proteins. Therefore, it is of interest to study the mechanism of degradation of oxidatively damaged proteins in synaptosomes. One way of oxidizing synaptosomal proteins in vitro is by incubating intact synaptosomes in the presence of an oxidizing agent. A problem with this approach is that it may also cause oxidative damage to the machinery required to recognize and degrade oxidized proteins. We have, therefore, introduced a fluorescent macromolecule into synaptosomes to assess the feasibility of using this technique to study how oxidized proteins are degraded and removed from synaptic terminals. Synaptosomes were subjected to electroporation in the presence of FITC labelled-dextran with an average molecular weight of 70000 (FD-70) and non-specific binding was determined by running parallel experiments in lysed synaptosomes. Following extensive washing, synaptosomes were assayed for the presence of intra-synaptosomal FD-70 by measuring fluorescence in a microplate fluorescence reader. Significant differences in fluorescence were found between intact and lysed synaptosomes with maximal uptake at 100 V/ 1500 microF (approx. 36 pmol/mg protein). To determine if membrane transport was compromised by electroporation, uptake of 3H-arginine was compared in control and electroporated synaptosomes. While untreated electroporated synaptosomes showed a loss of 22% in the ability to transport arginine, preincubation in the presence of 1 mM ATP resulted in a complete restoration of arginine transport. These results show that electroporation is a potentially useful technique for introducing a specific oxidized protein, into synaptic terminals so its metabolic fate can be examined.

Positioning during hemodynamic monitoring: evaluating the research.

Can pulmonary artery or right atrial pressures be obtained accurately when patients are in lateral positions? This review of the published research says probably not.

Backrest elevation and pulmonary artery pressures: research analysis.

Obtaining pulmonary artery pressure readings in the traditionally accepted flat, supine position can be uncomfortable for the patient and time consuming for the nurse. This analysis of published research suggests that some patients need not lie flat.

Effects of backrest elevation and position on pulmonary artery pressures.

This repeated measures study identified each subject's baseline range of fluctuation in pulmonary artery pressures and used this information to evaluate the effects of changes in backrest elevation (0 degrees and 30 degrees) and position (supine and lateral) on pulmonary artery pressures. Sixteen hemodynamically stable patients were studied. Six measures of pulmonary artery pressure were obtained with subjects in the supine (flat) position. Pulmonary artery systolic pressures fluctuated from 2-16 mm Hg, pulmonary artery diastolic pressures from 0-11 mm Hg, and pulmonary artery mean pressures from 0-13 mm Hg. Only pulmonary artery diastolic pressures were not altered in a statistically significant way with backrest elevation. Even though backrest elevation produced clinically significant changes (those exceeding the subject's baseline range of fluctuation) in some subjects, no other changes in parameters of hemodynamic status were observed. These results suggest that pulmonary artery pressures may be obtained with backrests elevated. Since position changes produced both statistically and clinically significant changes in all pulmonary artery pressures, the use of lateral positions during pulmonary artery pressure measurement is not recommended.