Chemical constituents isolated from Paeonia lactiflora roots and their neuroprotective activity against oxidative stress in vitro.

The methanolic extract of Paeonia lactiflora roots significantly protected primary cultures of rat cortical cells exposed to oxidative stress induced by H(2)O(2). Seven monoterpenes, paeonilactone-B (1), paeonilactone-C (2), paeoniflorigenone (3), benzoylpaeoniflorin (4), paeoniflorin (5), oxypaeoniflorin (6) and albiflorin (7), were isolated by bioactivity-guided fractionation and further separation using chromatographic techniques. Among them, compounds 2 and 4 significantly protected primary cultures of rat cortical cells against H(2)O(2)-induced neurotoxicity.

Blood harmane is correlated with cerebellar metabolism in essential tremor: a pilot study.

BACKGROUND: On proton magnetic resonance spectroscopic imaging ((1)H MRSI), there is a decrease in cerebellar N-acetylaspartate/total creatine (NAA/tCr) in essential tremor (ET), signifying cerebellar neuronal dysfunction or degeneration. Harmane, which is present in the human diet, is a potent tremor-producing neurotoxin. Blood harmane concentrations seem to be elevated in ET. OBJECTIVES: To assess in patients with ET whether blood harmane concentration is correlated with cerebellar NAA/tCR, a neuroimaging measure of neuronal dysfunction or degeneration. METHODS: Twelve patients with ET underwent (1)H MRSI. The major neuroanatomic structure of interest was the cerebellar cortex. Secondary regions were the central cerebellar white matter, cerebellar vermis, thalamus, and basal ganglia. Blood concentrations of harmane and another neurotoxin, lead, were also assessed. RESULTS: Mean +/- SD cerebellar NAA/tCR was 1.52 +/- 0.41. In a linear regression model that adjusted for age and gender, log blood harmane concentration was a predictor of cerebellar NAA/tCR (beta = -0.41, p = 0.009); every 1 g(-10)/mL unit increase in log blood harmane concentration was associated with a 0.41 unit decrease in cerebellar NAA/tCR. The association between blood harmane concentration and brain NAA/tCR only occurred in the cerebellar cortex; it was not observed in secondary brain regions of interest. Furthermore, the association was specific to harmane and not another neurotoxin, lead. CONCLUSION: This study provides additional support for the emerging link between harmane, a neurotoxin, and ET. Further studies are warranted to address whether cerebellar harmane concentrations are associated with cerebellar pathology in postmortem studies of the ET brain.

Effect of treatment with the dihydropyridine-type calcium antagonist darodipine (PY 108-068) on the expression of neurofilament protein immunoreactivity in the cerebellar cortex of aged rats.

The effect of long-term treatment with the dihydropyridine-type Ca2+ antagonist darodipine (PY 108-068) on the expression of neurofilament (NF) protein (200 kDa-NF subunit) immunoreactivity in the cerebellar cortex of aged male Wistar rats was assessed using immunohistochemical techniques associated with image analysis. In 12-month-old rats (adult) used as reference animals, 200 kDa-NF subunit immunoreactivity was observed primarily in axons of basket neurons localized in the molecular layer and surrounding the cell body of Purkinje neurons. A specific immunoreactivity was also found in the initial segment of Purkinje neuron axons, and in axons of the white matter of the cerebellar cortex. In 24-month-old rats (aged) a significant decrease in the area occupied by immunoreactive structures was noticeable in comparison with adult animals. A 6-month treatment (from the 18th to the 24th month of life) with an oral daily dose of 10 mg/kg of darodipine restored in part the expression of 200 kDa-NF subunit immunoreactivity in the cerebellar cortex. These data indicate that treatment with the dihydropyridine-type Ca2+ channel blocker darodipine is able to counter in part the age-related loss in the expression of NF protein in the rat cerebellar cortex. This suggests that darodipine may reduce neuronal cytoskeletal changes occurring in aging and in neurodegenerative disorders.

A rat brain cDNA encodes enzymatically active GABA transaminase and provides a molecular probe for GABA-catabolizing cells.

cDNAs encoding gamma-aminobutyric acid aminotransferase (GABA-T) were isolated from a lambda ZAP rat hippocampal cDNA expression library by two independent cloning methods, immunological screening with an antimouse GABA-T antibody and plaque hybridization with a GABA-T cDNA probe derived by polymerase chain reaction. We have produced enzymatically active GABA-T from a rat brain cDNA containing the full-length GABA-T coding region. Our rat brain GABA-T cDNAs hybridize to mRNAs in brain and peripheral tissues, including liver, kidney, and testis. We have also detected GABA-T mRNA in GABAergic cells of rat cerebellar cortex by in situ hybridization. Our rat brain GABA-T probe hybridizes to Purkinje, basket, stellate, and Golgi II cells, the same GABAergic neurons previously shown to contain glutamate decarboxylase GAD65 and GAD67.

Age-related changes in the relative abundance of NMDAR1 mRNA spliced variants in the rat brain.

We have investigated the age-dependent profile of two groups of NMDAR1 mRNA isoforms, the NR1(0)XX and the NR1(1)XX, which are characterized by the absence and the presence, respectively, of an N-terminal positioned 21 amino acid insert. mRNAs of the two spliced variants were investigated at different ages in discrete rat brain areas by means of the reverse transcription-polymerase chain reaction. The existence of regional variations was confirmed as well as a region-specific pattern of NR1(0)XX/1XX ratio and its age-dependent changes. At 6 months, the ratio was > 3 in prosencephalic structures and < 1 in metencephalic regions. The greatest age-related changes were found in the cerebellum that switched from a maximal ratio of 5.1 +/- 0.4 at day 6 through a progressive decay down to the value of 0.3 +/- 0.04 at 24 months. Age-dependent changes of the different NR1 spliced variant mRNAs are relevant to understand possible regulatory mechanisms of the pharmacological properties and functions of different NMDAR subtypes.

Quantitative scanning transmission electron microscopy of ultrathin cryosections: subcellular organelles in rapidly frozen liver and cerebellar cortex.

Freeze-dried, ultrathin cryosections of directly frozen mouse liver and brain have been prepared and characterized by low-dose dark-field scanning transmission electron microscopy (STEM). These improved cryosections gave images comparable to those from conventional plastic sections. They were thin enough (< < 1.0 elastic mean free path) to use established dark-field techniques, modified for thickness-dependent nonlinearities, to measure the dry mass fraction of individual organelles, and hence to deduce their water content. Digital STEM imaging in combination with electron and X-ray spectroscopy has important biological applications, as illustrated by studies on calcium regulation in Purkinje neurons. Calcium concentrations per unit dry weight of dendritic compartments were determined by the peak/continuum method of energy-dispersive X-ray spectroscopy (EDXS), which necessarily overstates elemental concentrations because of beam-induced mass loss. The dry mass content of organelles at low dose and the percentage of dry mass retained after analysis at high dose were as follows: mitochondria (46.0 g dry mass/100 g hydrated mass, 67% mass retained); endoplasmic reticulum (27.9 g/100 g, 57%); and cytoplasm (16.3 g/100 g, 41%). These values were used to correct elemental concentrations for mass loss. Results indicated that the major calcium storage organelle in Purkinje cell dendrites is the endoplasmic reticulum, of which there are two types distinguished by their levels of calcium. Parallel electron energy loss spectroscopy of dendritic organelles corroborated EDXS measurements, with an improved sensitivity that indicates the feasibility of quantitative calcium mapping.

Distribution of parvalbumin immunoreactivity in the human brain.

The cellular distribution of parvalbumin-like immunoreactivity (PA-LI) in the human brain was investigated by peroxidase-antiperoxidase methods using antiserum to rat skeletal muscle parvalbumin. PA-LI was present in non-pyramidal neurons of the cerebral cortices, stellate cells, basket cells and Purkinje cells in cerebellar cortices, and neurons of some nuclei in human brain stem; the distribution of PA-LI in human brain was very similar to that in rat brain. These results indicate that PA-LI is widely distributed in a specific subpopulation of neurons of the human brain.