Modified calcium accumulation after controlled cortical impact under cyclosporin A treatment: a 45Ca autoradiographic study.

OBJECTIVE: As a neuroprotective drug, cyclosporin A (CsA) has been subject of multiple experimental works in traumatic brain injury (TBI) research. It is well known that CsA inhibits calcium (Ca2+) induced mitochondrial permeability transition (mPT). The aim of this study was to investigate the influence of CsA on the alteration of Ca2+ homeostasis after experimental brain injury. METHODS: Sprague-Dawley male rats (n = 36) with a mean weight of 330 g (280-350 g) were general anesthetized with isofluran through gas mask. The anesthetized animals (n = 24) were subjected to a controlled cortical impact (CCI) over the left parietotemporal cortex using round-tip impounder with a 5 mm diameter at a velocity of approximately 3.7 m/s and a penetration depth of 2 mm. The sham group (n = 12) underwent anesthesia and craniotomy without CCI. In the CCI groups, CsA (n = 12) or vehicle (n = 12) was administered 15 minutes post-injury with a subsequent i.p. injection after 24 hours. Thirty-three hours after injury or sham craniotomy, 45calcium (45Ca) suspended in physiologic saline solution was injected in the left femoral vein. Five hours after isotope administration, animals were killed and the brain was quickly removed and placed in powdered dry ice. Coronal plane sections (20 microm thick) taken every 400 microm from the frontal cortex through the occipital cortex, were exposed to cyclotron films for 14 days at -18 degrees C. Relative optical density was utilized to provide a relative measure of 45Ca accumulation within seven different structures. RESULTS: The difference of 45Ca accumulation (measured by relative optical density) in the CsA group was greater by 30-70% in the following structures compared to vehicle treated traumatized animals: temporal cortex, CA1, anteromedial and posteromedial thalamus (p < 0.05). CONCLUSION: Post-traumatic 45Ca accumulation is modified under CsA. The crucial neuroprotective effect of CsA might be unrelated to a reduction of post-traumatic Ca2+ accumulation, especially with regard to the importance of Ca2+ as an intracellular messenger governing a large number of cellular functions.

The potential of differential mobility analysis coupled to MS for the study of very large singly and multiply charged proteins and protein complexes in the gas phase.

As previously demonstrated by the technique of gas-phase electrophoretic mobility molecular analyzer (GEMMA) introduced by Kaufman and colleagues, differential mobility analysis (DMA) of charge-reduced electrospray ions in the gas phase is a useful complement to MS for studying large proteins and their weakly bound complexes. Several limitations of GEMMA, the solutions for which have the potential to greatly improve its performance, are discussed here, including DMA resolution and transmission. A quantitative theory of charge reduction kinetics for dried multiply charged globular proteins at atmospheric pressures is also presented, showing that the charge reduction time must be carefully chosen to maximize a singly charged ion signal, while avoiding survival of contaminating multiply charged species. Because charge reduction limits the range of masses analyzable by MS, we also consider the potential of a parallel-plate DMA coupled in series to an MS for DMA-MS studies without charge reduction.

Aerosol size standards in the nanometer size range II. Narrow size distributions of polystyrene 3-11 nm in diameter.

A technique for generating charged aerosols of polystyrene (pSty) with narrow size distributions has been developed. It is based on electrospraying commercial narrow mass standards of pSty dissolved in l-methyl-2-pyrrolidone (NMP) seeded with the newly synthesized salt dimethyl ammonium formate. This salt imparts a much larger electrical conductivity than previously known NMP electrolytes, leading to higher quality sprays with greatly reduced attachment of impurities. Controlling the solute concentration enables forming polystyrene particles containing from one up to more than ten single polystyrene molecules, whereby 4 mass standards with molecular weights from 9200 up to 96,000 g/mol yield particles covering densely the diameter range from 3 to 11 nm. Combined mobility and mass measurement with a differential mobility analyzer and a mass spectrometer in tandem are carried out with a pSty sample 9200 amu in molecular weight. They fix directly the mass versus mobility relation near 9200 amu, and indirectly for the other standards and their clusters. The apparent particle density resulting from mobility versus mass data agrees with the bulk density of the polymer, indicating that the particles are dense and spherical. Although these standards have been studied only in gaseous suspension, their injection in liquids such as water where pSty is insoluble should keep them spherical.

Mass analysis of water-soluble polymers by mobility measurement of charge-reduced ions generated by electrosprays.

Aqueous solutions of poly(ethylene glycol) (PEG) in a 10 mM ammonium acetate buffer are electrosprayed, and the maximum charge state on the resulting gas-phase ions is reduced to unity using a radioactive source. The mobility distribution of these charged particles is then measured in air in a differential mobility analyzer of unusually high resolution. The relation Z(m) between the mobility Z of a polymer molecule and its mass m is determined by means of narrowly distributed PEG mass standards. The molecular weight range of available standards is extended by generating clusters containing from one up to six molecules of the primary PEG standard. The mass at the peak of the distribution of the lowest standard (PEG-4k) is determined by MALDI mass spectrometry and agrees with the manufacturer's value and previous MALDI literature data. The masses for the 50K and 120K standards are found to differ by 8.6 and 6.6%, respectively, from the manufacturer's value. Using known relationships, the particle diameter d of the ions is calculated from the measured mobility. Plots of d versus m(1/3) give straight lines over the full mass range studied (4000-700 000 Da, particle diameter from 3 to 12 nm), indicating that these PEG particles are indeed spherical and have a density rho independent of size. The slope of the d versus m(1/3) curve provides a density rho = 1.25 g/cm(3), close to the known bulk density, rho(PEG) = 1.21 g/cm(3).