Effects of ovariectomy and estradiol replacement on the binding of 125I-neurotensin in rat suprachiasmatic nucleus

Distribution and density of specific high-affinity 125I-neurotensin-binding sites were examined by light microscopic radioautography in the suprachiasmatic nucleus (SCN) of normal cycling, ovariectomized, or ovariectomized and estradiol-implanted female rats. In all three experimental groups, intense 125I-neurotensin labeling was detected within the ventrolateral component of the SCN. Whereas the topographic distribution and spread of the label was similar between each group, the density of the label was significantly higher (mean increase: 122%) in ovariectomized than in normally cycling females within the rostral third of the SCN. This effect was no longer apparent in females chronically implanted with estradiol at the time of gonadectomy. These results indicate that plasma gonadal steroids may regulate neurotensin receptors in the rat SCN. It is suggested that this mechanism might be implicated in the feedback control of gonadotropin and prolactin secretion.     

Prognostic factors in dilated cardiomyopathy

Thirty-eight patients, 27 men and 11 women, mean age 49 +/- 12 yr, suffering from dilated cardiomyopathy were included in this prospective study of prognostic factors. Twenty-two subjects admitted heavy alcohol consumption (greater than 80 g/d, for at least 10 yr). The survival rates for 2, 5 and 7 yr were 87, 35 and less than 20%, respectively. Prognostic factors were evaluated in 34 (22 alcoholic and 12 non-alcoholic) patients: 10 improved or were cured, as determined clinically, radiologically and echocardiographically (group 1); 24 deteriorated, 13 of whom died (group 2). Statistically significant values for group 1 versus group 2 were: echocardiographic left ventricular (LV) end diastolic diameter (3.50 +/- 0.80 vs 4.02 +/- 0.5 cm/m2, p less than 0.026) and LV end systolic diameter (2.97 +/- 0.65 vs 3.43 +/- 0.53 cm/m2, p less than 0.02); hemodynamic LV systolic pressure (113.9 +/- 15 vs 101.82 +/- 17.36 mm Hg, p = 0.045); angiographic LV end diastolic volume (137.8 +/- 57.44 vs 177.85 +/- 55.45 ml/m2, p = 0.057), LV end systolic volume 93 +/- 53.5 s 139.28 +/- 48.99 ml/m2, p less than or equal to 0.036), ejection fraction 0.36 +/- 0.16 vs 0.22 +/- 0.7%, p less than or equal to 0.006) and velocity of fiber shortening (0.79 +/- 0.53 vs 0.36 +/- 0.3 circ/s, p = 0.036). Other electrical, radiological and hemodynamic parameters were not significant and alcohol consumption did not influence the prognosis. The severity of LV functional impairment appears to be the major determinant of a poor prognosis and reduced survival in idiopathic and alcoholic cardiomyopathies.     

Lysosomes Contribute to Anomalous Pharmacokinetic Behavior of Melanocortin-4 Receptor Agonists

Purpose A series of melanocortin-4 receptor (MC4R) agonists, developed for use as anti-obesity agents, were found to have unusual pharmacokinetic behavior arising from excessive retention in the liver, with nearly undetectable levels in plasma following oral administration in mice. This work investigates the molecular basis of the prolonged liver retention that provided a rational basis for the design of an analog with improved behavior. Materials and Methods The livers of mice were harvested and techniques were utilized to fractionate them into pools differentially enriched in organelles. The distribution of organelles in the fractions was determined using organelle-specific enzymatic assays. Livers from mice dosed with drug were fractionated and comparisons with organelle distributions assisted in determining the subcellular localization of the drug. Further analysis in cell culture systems was used to confirm results from liver fractionation studies and also allowed for more extensive evaluations to examine the mechanism for organelle compartmentalization Results Fractionation of livers following oral administration of the agonist showed sequestration in lysosomes. Subsequent evaluations in a cell culture system confirmed this finding. Agents used to disrupt acidification of lysosomes led to decreased lysosomal accumulation of the drug, which implicated a pH-partitioning type sequestration mechanism. These findings led to the rational synthesis of an analog of the parent compound with properties that reduced lysosomal sequestration. When this compound was examined in mice, the liver retention was found to be greatly reduced and plasma levels were significantly elevated relative to the parent compound. Conclusions Weakly basic drugs with optimal physicochemical properties can be extensively sequestered into lysosomes according to a pH-partitioning type mechanism. When administered orally in animals, this particular sequestration event can manifest itself in long term retention in the liver and negligible levels in blood. This work revealed the mechanism for liver retention and provided a rational platform for the design of a new analog with decreased liver accumulation and better opportunity for pharmacokinetic analysis and therapeutic activity.     

Sympathetic neuroaxonal dystrophy in the aged rat pineal gland

Dysfunction of circadian melatonin production by the pineal gland in aged humans and rats is thought to reflect the functional loss of its sympathetic innervation. Our ultrastructural neuropathologic studies of the sympathetic innervation of the pineal gland of aged (24 months old) Fischer-344 and Sprague-Dawley rats showed loss of nerve terminals as well as the development of neuroaxonal dystrophy (NAD), an ultrastructurally distinctive distal axonopathy, far in excess of that in young control rats. Immunolocalization of tyrosine hydroxylase confirmed the age-related loss of normal noradrenergic innervation and development of NAD. NAD was more frequent in aged female rats compared to males and was particularly severe in aged female Sprague-Dawley rats compared to Fischer-344 rats. Pineal NGF content was significantly increased or unchanged in female and male aged Fischer-344 rats, respectively, compared to young controls. The rat pineal is a sensitive experimental model for the quantitative ultrastructural examination of age-related neuropathological changes in nerve terminals of postganglionic noradrenergic sympathetic axons, changes which may reflect similar changes in the diffusely distributed sympathetic innervation of other targeted endorgans.     

Leukocyte migration in adipose tissue of mice null for ICAM-1 and Mac-1 adhesion receptors

OBJECTIVE: To determine whether the leukocyte adhesion receptors ICAM-1 and Mac-1, regulators of immune cell migration, have an intrinsic role within adipose tissue by 1) analyzing the expression of ICAM-1 in adipose tissue, 2) identifying leukocyte populations within adipose tissue, and 3) determining whether ICAM-1 and Mac-1 mutant mice exhibit abnormal numbers of adipose tissue leukocytes. RESEARCH METHODS AND PROCEDURES: Wild-type, ICAM-1(-/-), and Mac-1(-/-) mice were fed a long-term high-fat diet. ICAM-1 expression was analyzed by Northern blot and immunohistochemistry. Leukocytes within adipose tissue were identified by immunohistochemistry and flow cytometry. RESULTS: ICAM-1 was expressed in adipose tissue and localized to the vascular endothelium. Macrophages and lymphocytes were prevalent within the stromal-vascular cell fraction of adipose tissue, and gender-specific differences were observed, with adipose tissue from female mice containing significantly more macrophages than tissue from male mice. Numbers of leukocytes in ICAM-1(-/-) and Mac-1(-/-) mice were not different from wild-types, however, indicating that these adhesion receptors are not required for leukocyte migration into adipose tissue. DISCUSSION: Our results documented leukocyte populations within adipose tissue, which may be involved in the development of heightened inflammation that is characteristic of obesity.     

Functional polymorphisms in the human β4 subunit of nicotinic acetylcholine receptors

Human nicotinic acetylcholine receptor (nAChR) polymorphisms occur in different ethnic populations and may result in differences in nAChR ion channel properties. We have identified four nAChR beta 4 subunit (4) nucleotide variants: 392CT, 526CT, 538AG, and 1519AG. Their corresponding amino acid substitutions are: Thr to Ile at codon 91 (T91I), Arg to Trp at codon 136 (R136W), Ser to Gly at codon 140 (S140G), and Met to Val at codon 467 (M467V), respectively. The nAChR ion channel properties of these variants were studied and compared with the more-common (wild-type) allele as wild-types. The nAChRs (44 channels) were expressed heterologously in Xenopus oocytes and studied using the two-electrode voltage clamp technique to reveal functional differences between the wild-type and the variants. The receptors containing the R136W and M467V mutations (or variants) had a higher sensitivity to acetylcholine and lower EC₅₀ than the wild-type. The T91I mutation had lower sensitivity to acetylcholine and the EC₅₀ was larger than in wild-type nAChRs. The S140G mutation had a dose-response relationship that was similar to the wild-type. The T91I, R136W, and M467V mutations (or variants) also showed a slightly greater degree of steady-state desensitization than the wild-type in response to a 30-min exposure to one tenth of their EC₅₀. The present results demonstrate that human 4 nAChR DNA polymorphisms result in functional changes, and suggest that certain individuals with those variants may be more or less sensitive to cholinergic drugs or to dysfunctions associated with nicotinic cholinergic systems.     

Sub-population of cultured hippocampal astrocytes expresses neuropeptide Y Y(1) receptors

The expression and pharmacological characterization of neuropeptide Y (NPY) receptors of the Y(1) subtype on cultured hippocampal neurons was reported using radioreceptor assays and immunohistochemical approaches (St-Pierre et al., 1998). The present study aimed to establish the presence of NPY Y(1) receptors on cultured hippocampal astrocytes using similar strategies. Immunocytochemical experiments were carried out using three antisera directed against distinct domains (amino acids sequence 185-203, 198-213 and 355-382) of the Y(1) receptor. Double-labeling experiments and confocal microscopy with these Y(1) receptor antisera demonstrated their recognition of the same sub-population (20%) of GFAP-positive astrocytes in culture. The immunostaining seen with all three Y(1) receptor antisera took the form of large irregular clusters distributed throughout cell bodies and processes. Further experiments using radioactive ligands confirmed the presence of NPY receptors on cultured hippocampal astrocytes. Emulsion receptor autoradiography using a newly developed ligand, [(125)I]GR231118 in the presence of PYY, hPP or BIBP3226 (1 microM), pharmacologically established the Y(1) nature of these receptors. Specific [(125)I]GR231118 binding was competed by PYY and the selective Y(1) antagonist BIBP3226 but not by hPP (a Y(4)/Y(5) agonist). Similar autoradiographic labeling patterns were obtained using [(125)I][Leu(31).Pro(34)]PYY (a Y(1)/Y(4)/Y(5) agonist) whereas [(125)I]PYY(3-36) (a Y(2)/Y(5) agonist) failed to generate any specific signal. Hence, rat cultured hippocampal astrocytes express a typical Y(1) receptor without evidence for the presence of Y(2), Y(4) or Y(5) subtypes. These data suggest a preferential regulation by NPY, acting via the Y(1) receptors, of astrocytic function.     

Respiratory protective devices: rates of medical clearance and causes for work restrictions

BACKGROUND: There are no published data on the outcomes and benefits of medical evaluations for the use of respiratory protective devices. We, therefore, conducted a retrospective database and chart review to assess the rates of medical clearance and causes for work restrictions at a Department of Energy complex. METHODS: All workers with work restrictions or denied clearance over a one-year period were identified and their medical records abstracted. RESULTS: Of the 5,569 workers who received medical evaluation, only 71 (1.3%) received limitations on respirator use documented in their medical record. Of the 65 workers with sufficient medical records for additional analysis, 9 of the 5,569 workers (0.2%) were denied medical clearance, while 56 workers (1.1%) received work restrictions. Pregnancy was the most common cause for denying medical clearance for respirator use. Lung disease, cardiovascular disease, and claustrophobia were the most common causes for work restrictions. Physical examination and spirometry added little to the detection of relevant medical conditions. CONCLUSIONS: We conclude that few workers fail medical clearance for respirator use or receive work restrictions. Data on adverse events from respirator use are needed to help design appropriate medical evaluations and uniform criteria for work restrictions or denial of medical clearance.