Biomedical subjects

C J Phelps

Publications and source records attributed to C J Phelps.

49 records · Page 3Linked to original sources

PubMed1987

Effect of chronic hyperprolactinemia on tuberoinfundibular dopaminergic neurons: histofluorescence in aged and in diethylstilbestrol-treated male rats.

Catecholamine histofluorescence was used to examine morphology of hypothalamic tuberoinfundibular dopaminergic (TIDA) median eminence-afferent neurons (areas A12 and A14) in male Fischer 344 rats bearing prolactin (PRL)-secreting tumors. These tumors were either spontaneous, discovered in rats aged 20-32 months, or were induced by chronic diethylstilbestrol (DES) treatment in younger animals. In the latter case, histofluorescence was examined in two groups of animals: (1) those which had been treated, beginning at 6 months of age, with 8 weeks of DES, and subsequently had had the treatment discontinued for 10 months, but which continued to exhibit hyperprolactinemia; (2) animals treated continuously with DES, for 50 or 70 days. Histofluorescence was induced using either the Falck-Hillarp, aluminum-formaldehyde ('ALFA') or the formaldehyde-glutaraldehyde ('FAGLU') technique and pituitaries were examined for PRL immunoreactivity, as permitted by the fixation method chosen for histofluorescence. For selected animals, pituitaries were enzymatically dissociated, maintained in primary culture, and were assessed for PRL cell quantification, intracellular PRL content, and in vitro release. Among animals treated chronically with DES, histofluorescence was assessed by the Falck-Hillarp technique, and counts of A12 and A14 perikarya were made in each hypothalamus. Median eminence zona externa fluorescence was diminished in aged rats with normal pituitaries, in aged rats bearing spontaneous tumors, and in rats treated continuously with DES, in comparison with fluorescence in younger rats with normal pituitaries or from which DES treatment had been with-drawn. Perikaryal fluorescence showed typical cell morphology and numbers in aged rats, aged rats bearing spontaneous tumors, and in rats from which DES had been withdrawn. In one animal, treated continuously for 70 days with DES, numbers of A12 and A14 perikarya decreased. In aged rats bearing spontaneous tumors, histofluorescence morphology indicated stimulatory effects on TIDA perikarya and terminals, including increased perikaryal and terminal fluorescence intensity. The morphological results support biochemical evidence of PRL stimulation of tuberoinfundibular dopaminergic neurons, and suggest that inhibitory effects of increased PRL secretion on these hypothalamic cells in DES-induced hyperprolactinemia reflect a direct and reversible effect of DES on these neurons.

Aging↗

PubMed1986

Immunocytochemical analysis of prolactin cells in the adult rat adenohypophysis: distribution and quantitation relative to sex and strain.

Prolactin immunocytochemistry was conducted 1) in serial sections of whole fixed pituitaries, for cell distribution analysis; and 2) on preparations of enzymatically dispersed anterior pituitary cells maintained for 1-4 days in vitro, for quantitation of the relative population of prolactin (PRL) cells. Both types of analysis were conducted on glands from young adult rats of both sexes in Long-Evans, Sprague-Dawley, and Fischer 344 strains. Cell quantitation results were compared with serum levels, intracellular content, and in vitro release of prolactin, for individual cell preparations. The results show that both PRL cell distribution and relative population numbers are similar between sexes in all three rat strains. Average percentages ranged from 25.9% to 32.1% in all young males and diestrous females. Prolactin-positive numbers of cells correlated positively with intracellular hormone content but not with serum or medium PRL levels. The prolactin cell population was significantly larger in aged Fischer 344 males but was not correlated with intracellular PRL levels. The prolactin cell population was heterogeneous in staining intensity and distribution in both sexes of all three strains.

Animals↗

PubMed1986

Pituitary lactotroph sedimentation profiles and in vitro secretory activity after ablation of the medial basal hypothalamus.

Pituitary cells from adult male rats subjected to chronic (6 and 10 weeks) medial hypothalamic ablation (MHA) were analyzed by unit gravity sedimentation to assess distribution of size and density of lactotrophs, and for subsequent in vitro prolactin (PRL) release in primary culture. Tinctorial staining (Herlant's tetrachrome) showed that initial preparations of cells from MHA rats were small and relatively undifferentiated. MHA cells did not sediment as far into the gradient as did cells from intact control pituitaries. Intracellular PRL content was lower in all gradient fractions of MHA cells. At 6 weeks after surgery, peak recovery of PRL was also in the upper portions of the gradient. In the 10-weeks group, however, peak PRL recovery from MHA cells was in a population that sedimented further, but more restrictedly, in comparison with control cells. At both postsurgical intervals, the majority of tinctorially or immunocytochemically identified lactotrophs from lesioned rats were lower in the gradient, indicating enlarged and denser cells. Relative numbers of lactotrophs (per pituitary) were increased 10 weeks after MHA. In vitro PRL release, over a maximum of 21 days culture, was comparable for cells from MHA rats and intact controls, according to daily per cell secretion rates and "production index" (hormone released/initial hormone content). By comparison, luteinizing hormone (LH) release was suppressed in culture compared to intact controls, and LH was recovered from gradient fractions of smaller cells. The results indicate that chronic removal of hypothalamic influence results in gradual prolactin cell hypertrophy and decreased hormone retention and in relative increase in numbers.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1986

Characterization of toxins A and B of Clostridium difficile with monoclonal antibodies.

Two monoclonal antibodies (MAbs) were used to learn more about the structures of Clostridium difficile toxins A and B. One of the antibodies, the PCG-4 MAb, reacted specifically with toxin A. This MAb precipitated toxin A and neutralized the enterotoxic but not the cytotoxic activity of the toxin. The site to which the antibody bound was resistant to denaturation with sodium dodecyl sulfate; however, it was destroyed by N-bromosuccinimide. Immunoblot analysis with the PCG-4 MAb revealed the presence of a large number of bands in preparations of denatured toxin A, suggesting that toxin A exists as an aggregate of smaller components. The antibody was covalently coupled to Affi-Gel 10, and the gel was used to purify toxin A from the culture filtrate of a highly toxigenic strain of C. difficile by immunoaffinity chromatography. The second antibody, the G-2 MAb, cross-reacted with toxins A and B. The cross-reaction was confirmed by immunoblot analysis. These results show that toxins A and B share an epitope and suggest that they have a common subunit. The G-2 MAb did not neutralize or precipitate either toxin. The site to which the G-2 MAb bound was partially destroyed by sodium dodecyl sulfate and was resistant to oxidation with N-bromosuccinimide.

Antibodies, Monoclonal↗

PubMed1985

Hypothalamic catecholamine histofluorescence in dwarf mice.

Brains of growth hormone (GH)- and prolactin (PRL)-deficient Ames (df/df) and Snell (dw/dw) dwarf mice and normal mice of the same strains were examined for catecholamine (CA) histofluorescence, with particular emphasis upon the hypothalamic tuberoinfundibular (A12) (arcuate nucleus/median eminence) region, which plays a role in the regulation of both GH and PRL. Dwarfs and normal animals of both types also were treated with a drug regimen to deplete sequentially neuronal CA stores (reserpine), inhibit CA oxidation (nialamide) and load dopaminergic A12 cells with exogenous transmitter (norepinephrine), in order to test viability and axonal transport capacity of A12 neurons. In both types of dwarfs, compared with normals, fluorescence was markedly reduced in the zona externa of the median eminence, which is normally rich in terminals from A12 neurons. Fluorescence in the median eminence was particularly weak in Ames dwarfs, and A12 perikarya were difficult to discern in this group. Snell dwarfs showed reduced fluorescence of A12 perikarya when compared with the brightly fluorescent perikarya seen in normal mice. In supraoptic and paraventricular nuclei, and in the zona interna of the median eminence, CA fluorescence attributable to NE was comparable among dwarfs and normals; fluorescence of dopaminergic perikarya in substantia nigra was also unaffected in dwarfs. Exogenously administered NE effected enhanced fluorescence of A12 perikarya in normal mice and in Snell dwarfs; NE treatment in the Ames dwarf, however, failed to increase significantly the faint fluorescence of A12 cell bodies. The results indicate that dopaminergic A12 neurons in Snell dwarf mice are present and viable.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1985

Monoclonal and specific polyclonal antibodies for immunoassay of Clostridium difficile toxin A.

Monoclonal antibody, affinity-purified antibody, and monospecific antiserum against toxin A were produced. The monoclonal antibody was an immunoglobulin G2a kappa chain isotype that immunoprecipitated toxin A, as shown by crossed immunoelectrophoresis. These antibodies were compared by counterimmunoelectrophoresis, latex agglutination, and indirect enzyme-linked immunosorbent assay for their sensitivity in detecting toxin A. Our findings indicate that these antibodies may be useful as immunodiagnostic reagents for Clostridium difficile disease.

Animals↗

PubMed1984

Development of the catecholamine innervation of the supraoptic nucleus in the Brattleboro rat.

The ontogenetic development of the noradrenergic innervation of the supraoptic nucleus was studied in the Brattleboro rat at late postcoital and early postnatal ages. This genetic mutant offers a useful model for analysis of neuronal development because of the absence of a specific peptide component of identifiable target neurons and has been used presently to eliminate the possibility that such substances are essential for the establishment of normal connectivity during postnatal development. In this model, catecholamine varicosities were seen in juxtaposition to vasopressin-deficient perikarya during the initial phases of postnatal development, but these varicosities gradually decreased in number suggesting the possibility that the target neuron peptide, or some functional aspect of the neuron, may be necessary for the normal maintenance of this neuronal interaction.

Adrenergic Fibers↗

PubMed1984

Plasticity of catecholaminergic neurons in aged rat brain: reinnervation and functional recovery after axotomy.

Regenerative growth at the lesion site, reinnervation of a target nucleus and functional manifestations of recovery were studied in aged (20 and 30 months old) rats subjected to long-term transection of catecholaminergic (CA) fibers which contact and influence neurons of the supraoptic nucleus (SON). Small bilateral knife cuts were placed stereotaxically just caudal and medial to the SON. CA histofluorescence, induced by formaldehyde-glutaraldehyde (FAGLU) or aluminum-formaldehyde (ALFA) methods, was examined in hypothalamus at 2, 14, 21 and 60 days postsurgically. Water consumption, and urine volume and osmolality, were monitored presurgically, and through survival times. Subtotal CA denervation in the SON, and typical axonal transmitter "pile-up" at the lesion site, were evident two days after surgery. Among these degenerative profiles, which persisted for up to three weeks, fine-sized new fibers were apparent at the lesion, beginning between 2 and 14 days, and persisting throughout the period studied. At 21 days, and progressively thereafter, SON neurons were rimmed with fluorescent varicosites. Water consumption initially was depressed, but returned to presurgical mean levels by nine days. Urine volume returned to normal by 32 days. Urine osmolality showed a recovery by approximately three weeks. These functional parameters rebounded to levels higher than presurgical means among 20 month old, but not 30 month old, rats beyond 6 weeks survival, concurrent with a morphological hyperinnervation. The results reaffirm morphological regeneration, and support reinnervation and functional recovery, which extend considerably into the aging process.

Afferent Pathways↗

PubMed1983

Regeneration of central catecholamine fibers in young and aged rat brain.

Catecholamine histofluorescence patterns were examined in brains of young and aged rats, 1 to 14 days following neurosurgical transection of the medial forebrain bundle. At all ages examined, two phenomena were observed: degeneration of nerve fibers and vigorous regrowth of catecholamine-containing fibers in the lesion site. Regenerated catecholamine fibers invaded the area of scarred tissue. This invasion of the scarred area implies that the robust plasticity of catecholaminergic pathways, known to exist in young animals, persists in aged brain.

Aging↗

PubMedDate not supplied

Isolated deficiency of immunocytochemically detected somatostatin in Snell dwarf, but not in "little," mice.

Immunocytochemical staining of the neuropeptide somatostatin was evaluated in the brains of two growth hormone-deficient mouse mutants, Snell dwarf (dw/dw), and "little" (lit/lit). In Snell dwarf mice, in which GH is undetectable, an isolated somatostatin deficiency was observed in hypothalamic median eminence and in anterior periventricular hypothalamic neurons which are afferent to median eminence, compared to somatostatin staining in normal mice of the same strain (DW/?). Somatostatin staining in all other CNS areas in dwarfs was identical to that in normal mice. In contrast, in little mice, which exhibit 5-10% of normal GH levels, somatostatin expression was comparable between mutants and normal mice (LIT/?) in all CNS areas, including median eminence-afferent neurons in anterior periventricular hypothalamus. The results suggest that expression of somatostatin in hypophysiotropic CNS is dependent upon minimal pituitary GH secretion.

Animals↗

PubMedDate not supplied

Median eminence-afferent vasoactive intestinal peptide (VIP) neurons in the hypothalamus: localization by simultaneous tract tracing and immunocytochemistry.

Retrograde tract tracing and immunocytochemistry were used to investigate the CNS source of the VIP that is present in high concentrations in the hypophysial portal blood and has been shown to have a stimulatory effect on pituitary prolactin secretion. Fluoro-gold (FG), which enters the CNS through areas devoid of the blood-brain barrier, such as median eminence, was injected peripherally. Brain sections from FG-treated animals were immunostained for VIP. A small population of VIP-containing cell bodies in the parvocellular and periventricular parts of the paraventricular nucleus (PVN) was also labeled with FG. Vasoactive intestinal peptide-immunoreactive perikarya not labeled with FG were also observed in the PVN, as well as FG-labeled cells that did not contain VIP. The results suggest that some VIP-producing neurons in the PVN project to the median eminence and are, therefore, functionally related to pituitary regulation; the function of other VIP neurons in the PVN is unknown.

Animals↗

PubMedDate not supplied

Vasopressin in aged rats: longitudinal studies of vasopressin excretion in Sprague-Dawley and Fischer 344 strains.

In order to provide physiological baseline values for future experimental procedures, indices of vasopressin secretion were assessed in male Sprague-Dawley (SD) and Fischer 344 (F344) rats at 3 and 20 months of age. Daily water intake, urine volume, urine osmolality, and urinary vasopressin excretion were monitored in SD rats for 30 days, and in F344 rats for 60 days. In the SD strain, daily water and urine volumes, expressed as ml/24 hr/100 g b.wt., were consistently lower in aged animals, as was a calculation of water balance (water intake-urine output volumes/24 hr). Although mean VP concentration in urine appeared higher in aged rats (33.9 +/- 20.4 pg/ml) than in young (16.3 +/- 7.7 pg/ml), total daily VP excretion was comparable for both ages when expressed as a function of body weight [80.6 +/- 37.3 pg for 3 months old (m.o.) and 81.9 +/- 47.2 pg/24 hr/100 g b.wt. for 3 and 20 m.o. respectively]. Young and old F344 males showed comparable daily drinking and urine volumes, and water balance, during two months of monitoring, but VP excretion was lower (p less than 0.025) in aged rats (83.8 +/- 19.0 pg/24 hr/100 g b.wt.) than in 3 m.o. rats (213.0 +/- 48.1 pg/24 hr/100 g b.wt.). Urine VP concentration was comparable (69.6 +/- 20.6 for 3 m.o.; 59.8 +/- 25.6 pg/ml for 20 m.o.). Mean urine osmolality was not significantly different among groups. Urine osmolality and daily urine volumes showed a significant correlation with daily VP excretion among young, but not aged, rats of both strains.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗

PubMedDate not supplied

Effect of surgical deafferentation of the hypothalamic supraoptic nucleus on urinary vasopressin levels in twenty-month-old rats.

Indices of water balance (daily water consumption, urine volume, and urine osmolality), and vasopressin (VP) urinary excretion were measured in 3-month-old (m.o.) and 20 m.o. F344 rats after neurosurgical lesion of noradrenergic afferents to the supraoptic nucleus (SON) in the medial forebrain bundle (mfb), in order to assess whether morphological denervation (21) was functionally effective. Lesion of the mfb was accomplished using a mechanical knife cut placed stereotaxically medial and caudal to SON. The experiment was performed three times. Morphological evaluation consistently indicated a marked depletion of noradrenergic afferents in SON after lesion placement. The effect of the lesion on daily excreted VP was assessed by paired t-test, using means of equal numbers of urinary VP measurements representing 13-15 days each pre- and postsurgically for each animal. The postsurgical interval for assessing the functional effect of the lesion was chosen based upon morphological evidence for mfb regeneration at 14 days (21). Three m.o. rats showed comparable urinary VP levels before (mean = 328.4 +/- 97.3 pg/24 hr/100 g b.wt., for 10 rats for 2 weeks) and after (354.7 +/- 63.0 pg/24 hr/100 g b.wt.) lesion placement. In 20 m.o. rats, overall mean postsurgical VP excretion was 199.4 +/- 44.8 pg for 15 rats for 2 weeks, compared with a presurgical mean of 343.2 +/- 86.2 pg/24 hr/100 g b.wt. Mean urine VP was lower in 20 m.o. rats in each of three experiments; VP levels after lesion in 3 m.o. rats was lower in 2, but higher in one experiment; mean differences were not statistically significant for treatment (lesion) effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Age Factors↗