Ultrastructural observations of the cholinergic neuron in the rat striatum as identified by acetylcholinesterase pharmacohistochemistry

In order to examine the ultrastructural features of the cholinergic neuron in the striatum (caudatoputamen) of the rat, cytochemistry for acetylcholinesterase was conducted 2–12 h after intramuscular injection of the irreversible acetylcholinesterase inhibitor diisopropylphosphorofluoridate. Light microscopic examination of Epon sections reacted for acetylcholinesterase showed that only large-sized cells in the striatum (25–35 μm in the long axis) were stained intensely. In the case of longer survival periods (10–12 h), some lightly stained cells (medium-sized) were seen dispersed amongst the large acetylcholinesterase-rich cells. Electron microscopic observations were made on ultrathin sections of selected large acetylcholinesterase-rich neurons that were first studied by light microscopy. The nucleus of these cells has an eccentric position and possesses several indentations of the nuclear envelope. The cytoplasm contains abundant organelles, many exhibiting features unique to this cell type. Many stacks of granular endoplasmic reticulum, arranged in a parallel manner and forming typical Nissl bodies, were observed in the periphery of the perikarya, and many distinct golgi complexes were seen in the perinuclear zone. At all post-diisopropylphosphorofluoridate survival times, heavy deposits of acetylcholinesterase reaction product were found within the perikarya of this cell type, for the most part within the cisternae of the granular endoplasmic reticulum. At the longer post-diisopropylphosphorofluoridate survival times, reaction product within the cytoplasm was very dense and appeared to have reached a maximum level. At these times reaction product also appeared in the secondary and tertiary dendritic branches of the large-sized neurons.

Of the other cell types in the striatum, two types of medium-sized cells displayed a light deposit of reaction product in their perikarya, but this was observed only at longer recovery times (8–12 h). The majority of cells in the striatum lacked reaction particles. Throughout the early post-diisopropylphosphorofluoridate period, the recovery of enzyme activity in the neuropil was moderate compared to that seen within cell bodies.

These findings indicate that the large-sized neuron is the only striatal structure that shows rapid regeneration of acetylcholinesterase activity during the early recovery phase after diisopropylphosphorofluoridate administration. Previous studies have indicated that this type of neuron represents the cholinergic interneuron of the striatum. The present results indicate that under appropriate conditions acetylcholinesterase pharmacohistochemistry can be utilized to determine the ultrastructural features of central cholinergic neurons.     

The chitosan prepared from crab tendon I: the characterization and the mechanical properties

Crystalline chitosan was prepared from crab tendon consisting mainly of chitin, including various proteins and calcium phosphates. The crab tendon has high mechanical properties due to its aligned molecular structure. Crab tendon components, i.e. proteins and calcium phosphates, were removed by deacetyl treatment using 50wt% NaOH aqueous solution at 100 degrees C, and a subsequent ethanol treatment. As judged from microscopic observations using an optical polarizer, the treated chitosan remained intact regarding its aligned molecular structure, and had a high tensile strength of 67.9+/-11.4MPa. The tensile strength was further enhanced to 235+/-30MPa by a thermal treatment at 120 degrees C, corresponding to the formation of the intermolecular hydrogen bonds.     

Contrasting effects of hydroxyurea on cell growth and reduction in Epstein-Barr virus genomes in EBV-infected epithelioid cell lines vs Burkitt's lymphoma cell lines

Eliminating Epstein-Barr virus (EBV) genomes from infected cells is an intriguing theoretical strategy in therapy for EBV-associated malignant diseases. Respective patterns were characterized for hydroxyurea (HU)-promoted loss of EBV genomes from EBV-infected epithelioid cell lines derived from the noncancerous portion of gastric carcinoma tissues and Burkitt's lymphoma (BL) cell lines. Epithelioid cell lines GT38 and PN were less sensitivity to HU than BL cell lines Akata, Raji, and Daudi in terms of cell growth inhibition and cell cycle arrest. On passage in medium with 50 microM HU, the fraction of EBV nuclear antigen (EBNA)-positive cells was reduced substantially in the BL cell lines, but only slightly in the epithelioid cell lines. EBV DNA was reduced in Akata, Raji, and Daudi cells upon passage in 50 microM HU by 95%, 70%, and 50%, respectively, but by only 10% in GT38 cells, in which EBV DNA reduction was enhanced at increased concentrations of HU. This indicates that EBV genome is more easily lost from BL cell lines than from epithelioid cell lines upon culturing in HU. These findings support the view that the elimination of EBV could be therapeutically effective in EBV-associated BL by HU.     

Chemoprevention of heterocyclic amine-induced mammary carcinogenesis in rats

2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is one of the most prevalent carcinogenic heterocyclic amines in the environment, targeting the colon, prostate, pancreas, and mammary gland in rodents. Chemopreventive effects of synthetic and naturally occurring compounds on PhIP-induced rat mammary carcinogenesis were investigated in a series of experiments. In a PhIP feeding model, groups of 20-21 female F344 rats each, were treated with 0.02% PhIP alone or PhIP plus 0.5% 1-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ), 1% green tea catechins, 1% alpha-tocopherol, 0.1% ellagic acid, or 1% chlorophyllin, each in the diet, or 0.1% caffeine in drinking water for 52 weeks. To assess the mechanism of HTHQ and caffeine inhibition of PhIP-induced carcinogenesis, effects of these compound on the in vitro metabolic activation of PhIP were examined in the presence of S9 mix. In the next series of experiments, the PhIP intragastric dose model was applied to allow separate investigation of the effects of chemicals during the initiation and postinitiation periods. In these experiments, female Sprague-Dawley rats were given eight intragastric doses of 100 mg/kg body weight during the first 4-8 weeks for initiation. Either during initiation or after initiation, or only after initiation, animals were treated with either corn or perilla oil at doses of 5 and 20%, conjugated fatty acid derived from safflower oil (CFA-S) or perilla oil (CFA-P) at a dose of 1%, arctiin at doses of 0.02 and 0.2% in the diet, or sodium nitrite (NaNO(2)) at a dose of 0.2% in drinking water. In the PhIP feeding model, administration of PhIP alone for 52 weeks induced adenocarcinomas in 40% of rats, but the incidence was remarkably reduced to 5% by the simultaneous treatment with 0.5% HTHQ, a strong lipophilic phenolic antioxidant, or to 10% by 0.1% caffeine. Administration of 1% chlorophyllin exerted similar, albeit weaker, effects. alpha-Tocopherol at a dose of 0.5% only reduced the multiplicity of carcinomas, and 1% green tea catechins only the mean size of mammary tumors. In a metabolic activation study of PhIP, HTHQ and caffeine clearly inhibited the formation of metabolites. In the PhIP gastric dose model, among the naturally occurring compounds examined, a plant lignan arctiin, perilla oil, which contains a large amount of n-6 alpha-linolenic acid, and CFA-S or CFA-P inhibited mammary tumor development, particularly in the postinitiation period, although a clear dose response was not observed. Treatment with 0.2% NaNO(2) in the initiation period was found to lower the volume of mammary tumors. The present results indicate that a number of compounds may be candidate chemopreventive agents against PhIP-induced mammary carcinogenesis, acting through different mechanisms and depending on the stage of carcinogenesis.     

Different susceptibilities of long-term potentiations in CA3 and CA1 regions of guinea pig hippocampal slices to nootropic drugs

Effects of the nootropic drugs, piracetam and bifemelane, on long-term potentiation (LTP) of population spikes in the CA3 and CA1 regions of guinea pig hippocampal slices were investigated. Piracetam (10(-6) to 10(-4) M) and bifemelane (10(-8) to 10(-6) M) significantly augmented the LTP in the CA3 region. The effects of these drugs were inhibited by scopolamine (10(-6) M). However, the LTP in the CA1 region was not affected by piracetam and bifemelane even at highly effective concentrations (10(-5) and 10(-6) M, respectively). Thus, the LTP in the CA3 is more susceptible to nootropic drugs than in the CA1.