Inducible nitric oxide synthase stimulates dopaminergic neurodegeneration in the MPTP model of Parkinson disease.

MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) damages dopaminergic neurons as seen in Parkinson disease. Here we show that after administration of MPTP to mice, there was a robust gliosis in the substantia nigra pars compacta associated with significant upregulation of inducible nitric oxide synthase (iNOS). These changes preceded or paralleled MPTP-induced dopaminergic neurodegeneration. We also show that mutant mice lacking the iNOS gene were significantly more resistant to MPTP than their wild-type littermates. This study demonstrates that iNOS is important in the MPTP neurotoxic process and indicates that inhibitors of iNOS may provide protective benefit in the treatment of Parkinson disease.

A histological comparison of phase-partition fixation with fixation in aqueous solutions.

In phase-partition fixation, tissue is immersed in a non-aqueous solvent at equilibrium with an aqueous solution of a fixing agent to minimize osmotic effects. Preservation of morphology afforded by phase-partition fixation using formalin and glutaraldehyde and several organic solvents was compared to aqueous 10% neutral buffered formalin fixation for five tissues. It was shown that phase-partition fixation can provide excellent fixation for light microscopy if the proper combinations of fixatives and solvents are used.

Histochemistry and ultrastructure of the interstitium of the renal papilla in rats with hereditary diabetes insipidus (Brattleboro strain).

The Brattleboro strain of Long-Evans hooded rats has hereditary hypothalamic diabetes insipidus due to the inability to produce antidiuretic hormone. Animals homozygous for this autosomal recessive trait have extreme polyuria and polydipsia, whereas heterozygotes are less severely affected. Light and electron microscopy were used to study the interstitial tissue of the renal papilla of Brattleboro rats and normal Long-Evans rats. Staining with alcian blue or colloidal iron revealed that homozygous Brattleboro rats (DI) have greatly reduced quantities of glycosaminoglycans in the papillary interstitium. Heterozygotes showed staining similar but not identical to that of normal rats. The papillary interstitial cells of DI rats lacked the cytoplasmic processes seen in normal rats, and the normal relationship of these cells to the tubular elements of the papilla was absent. Electron microscopy revealed that the papillary interstitial cells of DI rats appeared less active than those of heterozygous or normal rats. In DI rats these cells displayed reduced numbers of lipid droplets and mitochondria, and the Golgi apparatus and rough endoplasmic reticulum were poorly developed. The altered ultrastructure of the papillary interstitial cells may be responsible for the reduction of interstitial glycosaminoglycans in DI rats. Glycosaminoglycans possess properties which may contribute to urinary concentration, It is suggested that the interstitial tissue of the renal papilla is influenced by antidiuretic hormone.

Effects of lithium on the structure of the rat kidney.

Light and electron microscopy were used to study the effects of a lithium-supplemented diet on renal structure in the rat. At the end of a 7-week experimental period serum lithium levels were 1.14 +/- 0.20 mM. Lesions consisting of groups of dilated tubules were found in the immediate vicinity of the interlobular arteries in all experimental animals. These tubules were identified as the connecting tubule or the initial portion of the collecting tubule. The epithelium of these tubules was generally flattened but was punctuated by markedly swollen epithelial cells. PAS-positive deposits found in both types of cells were identified as glycogen. Electron microscopy revealed considerable lithium-induced damage in the swollen cells including increased numbers of mitochondria, many of which were swollen or otherwise damaged, dilated cisternae of endoplasmic reticulum and vacuolization of the apical cytoplasm. The flattened cells of these tubules were similar to the dark or intercalated cells of normal collecting tubules. Some detachment of epithelial cells from their basement membrane was evident in these tubules. Damage was less severe in distal convoluted tubules. Lithium-induced changes were not observed in glomeruli, proximal tubules or ascending thick limbs of Henle. In medullary collecting tubules damage was less severe than in cortical collecting tubules, but detachment of epithelial cells was a common finding. The interstitial tissue of the papilla exhibited histochemical and ultrastructural changes consistent with lithium blockade of the action of antidiuretic hormone. The ultrastructural damage to cortical tubules is similar to that found in patients receiving therapeutic lithium for long periods of time. The anatomic sites of lithium-induced pathology correspond to the location of lithium-induced pathophysiology.

Human chromogranin A-like immunoreactivity in the Bergmann glia of the rat brain.

A mouse monoclonal antibody to human chromogranin A (LK2H10) selectively stains the Bergmann glia (Golgi epithelial cells) of the rat cerebellum, while rabbit antibodies to bovine or porcine chromogranin A do not. Chromogranin A immunostaining in the cell bodies and processes reveals the characteristic candelabra morphology seen in Golgi preparations of these cells. Immunostaining adjacent sections with antibodies to glial fibrillary acidic protein shows that a relatively small number of the glial cell processes in the molecular layer are chromogranin A positive. Recent reports indicate that the metabolites of chromogranin A play a regulatory role in endocrine and neuroendocrine systems but the function of chromogranins in neurons and glia is unknown.

A modification of the tannic acid-phosphomolybdic acid-dye stain for demonstrating myoepithelial cells in formalin fixed tissue.

A modified tannic acid-phosphomolybdic acid-dye procedure is used for staining myoepithelial cells in formalin fixed surgical and autopsy material. Paraffin sections are brought to water, mordanted for 1 hr in Bouin's fixative previously heated to 56 C, cooled while still in Bouin's, rinsed in tap water until sections are colorless, rinsed in distilled water, treated with 5% aqueous tannic acid 5-20 min, rinsed in distilled water 30 sec or less, treated with 1% aqueous phosphomolybdic acid 10-15 min, rinsed 30 sec in distilled water, rinsed in methanol, stained 1 hr in a saturated solution of amido black or phloxine B in 9:1 methanol:acetic acid, rinsed in 9:1 methanol:acetic acid, dehydrated, cleared and mounted. Myoepithelial cells of sweat, lacrimal, salivary, bronchial, and mammary glands are blue-green with amido black or pink with phloxine B. Fine processes of myoepithelial cells are well delineated. Background staining is minimal and the procedure is highly reproducible.

Acute neuropathological changes in the caudate nucleus caused by MPTP and methamphetamine: immunohistochemical studies.

Three days after the administration of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) or methamphetamine to mice, there is degeneration and disappearance of punctate tyrosine hydroxylase-containing synaptic endings in the caudate nucleus. The neuropil is occupied with longer, varicose, branching fibres, which appear to be preterminal fibres. An intense gliosis occurs. The sparsely-occurring glial cells, with profuse lightly-stained (by glial fibrillary acidic protein) processes which are primarily located near blood vessels, become transformed into more heavily-stained star-shaped cells with fewer but thicker processes. These cells are distributed throughout the caudate. Despite apparent differences in the mechanism by which MPTP and methamphetamine cause dopamine depletion, the neuropathological changes in the caudate induced by these substances are identical.

Suramin disrupts the gliotic response following a stab wound injury to the adult rat brain.

Reactive gliosis, observed in numerous pathological states, leads to the formation of a glial scar that is believed to impede axonal regeneration. Astrocyte reactivity can be initiated both in vitro and in vivo by various cytokines. Thus, the aim of this study was to investigate if suramin, a polysulfonated napthylurea that has been shown to inhibit the binding of many different cytokines to their cell surface receptors, could attenuate the glial response after brain injury. A single dose of suramin (5 microl, 75 microM) or saline vehicle was injected intracerebrally through the same needle used to make the stab wound at the time of lesioning. Suramin-treated animals showed an obvious reduction in several parameters of CNS inflammation: cellular proliferation, GFAP levels, and tenascin-C immunoreactivity were reduced in suramin-treated as compared to control animals at early time points. GFAP immunoreactivity was strikingly reduced at 3 days after injury, as confirmed by Western blot analysis. This reduction was transient, however, in that the difference in GFAP expression between suramin-treated and control animals was less apparent at 7 days and had disappeared by 30 days after injury. Likewise, fewer BrdU-positive cells were noted in treated versus control tissue at 1 and 3 days, but this difference was not significant by 7 days. Moreover, tenascin immunoreactivity was significantly diminished at 24 h as confirmed by Western blot analysis in suramin-treated lesion areas, which is analogous to our observations that suramin can antagonize tenascin expression by cultured astrocytes treated with bFGF. In addition, examination of the corpus callosum of saline-treated animals 30 days post-trauma revealed a disruption of the fiber tract within the lesion site, while suramin-treated animals displayed numerous fibers spanning the lesion. These results demonstrate that a single injection of suramin transiently inhibits the gliotic response, which may be sufficient to ameliorate subsequent tissue damage.