Biomedical subjects

K A Berghorn

Publications and source records attributed to K A Berghorn.

15 recordsLinked to original sources

PubMed2005

Developmental expression of the homeobox protein Distal-less 3 and its relationship to progesterone production in mouse placenta.

Distal-less 3 (Dlx3) is a homeobox factor that functions as a placental-specific transcriptional regulator. Dlx3 null mice (-/-) have compromised placental development and do not survive in utero past embryonic day (E) 9.5. The current studies were undertaken to examine the expression of Dlx3 in mouse placenta during gestation, and to determine whether Dlx3 was involved in placental progesterone production. Dlx3 was not detectable at E8.5 but was detected in E9.5 placenta with continuing but diminished expression through E15.5. Dlx3 immuno-localization was restricted to the labyrinth, was nuclear and was found in cytokeratin-positive cells. Previous studies in choriocarcinoma cell lines support the conclusion that Dlx3 is required for expression of 3'-hydroxysteroid dehydrogenase VI (3betaHSD VI), an obligate enzyme in the production of progesterone by trophoblast giant cells. In a rat trophoblast stem cell line (Rcho-1), Dlx3 expression was non-detectable in Rcho-1 cells induced to differ-entiate using mitogen withdrawal. In vitro progesterone production in placental cultures and 3betaHSD VI mRNA from Dlx3 (+/+), (+/-) and (-/-) mice were equivalent. In situ hybridization for 3betaHSD VI revealed mRNA expression restricted to trophoblast giants cells with no detectable expression in the labyrinth suggesting that Dlx3 and 3betaHSD VI were not colocalized within the placenta. These studies support the conclusion that Dlx3 protein expression is restricted to the labyrinth region of the murine placenta into late gestation and that Dlx3 does not appear to be expressed in trophoblast giant cells. Further, loss of Dlx3 was not correlated with synthesis of progesterone from E9.5 mouse placentas.

3-Hydroxysteroid Dehydrogenases↗

PubMed2003

Life before birth: effects of cortisol on future cardiovascular and metabolic function.

UNLABELLED: The concept of fetal programming is an area that is now under rigorous investigation in many laboratories throughout the world. We need to engender a fascination in all segments of society, not just pregnant women, about life in the womb. CONCLUSION: Everyone needs to understand that improving the condition of the fetus will have personal, social and economic benefits. The time has come to realize that, in a sense, it is not just women who are pregnant but it is the family and the whole of society.

Animals↗

PubMed2001

Suckling stimulus suppresses messenger RNA for tyrosine hydroxylase in arcuate neurons during lactation.

Tyrosine hydroxylase (TH) mRNA in tuberoinfundibular dopamine (TIDA) neurons is suppressed during lactation but rebounds upon pup removal. A time course of TH mRNA changes after pup removal revealed three phases: (1) a nuclear phase (evident 1.5 hours after pup removal, maximal at 3 hours) with TH mRNA appearing in 1 or 2 nuclear loci with little or no change in cytoplasmic mRNA; (2) a cytoplasmic phase (noted 6 hours after pup removal, peaked 12-24 hours) with a significant increase in total TH mRNA levels mainly in the cytoplasm; and (3) a stabilization phase (24-48 hours after pup removal) when nuclear signals were low and cytoplasmic RNA showed a slight decline with extension of RNA clusters into the cell dendrites. In rats whose pups could suckle only on one side, TH was up-regulated only on the side contralateral to nipple blockade. These data indicate that after suckling terminates, TH up-regulation is evident at 1.5 hours, but 6 hours is needed before the cells transport sufficient mRNA into the cytoplasm. The rapid signaling of TH up-regulation stems from the fact that the TIDA neurons respond to neural signals from termination of suckling.

Animals↗

PubMed2000

VIP innervation: sharp contrast in fetal sheep and baboon adrenal glands suggests differences in developmental regulation.

Immunocytochemical technique and light microscopy were used to ascertain the relationship between vasoactive intestinal polypeptide (VIP) and tyrosine hydroxylase in fetal sheep and fetal baboon adrenal cortices and medullae at 85% of gestation. VIP immunostaining was extremely robust in fetal sheep adrenal cortical neurofibers and cells while weak in fibers and nonexistent in cells of fetal baboon. Also, tyrosine hydroxylase-immunopositive cells, present throughout the adrenal cortices of both fetal sheep and baboons, were heavily innervated by VIP-immunoreactive neurofibers in fetal sheep, but not in fetal baboons. Adrenal cortical VIP-immunopositive fibers occurred in greater (P<0.05) frequency in fetal sheep than in fetal baboons (14.82+/-3.10 vs. 0.84+/-0.26 fibers/field), were larger in diameter (2.93+/-0.34 vs. 0.93+/-0.07 microm) and ran for longer distances in the plane of section (127.85+/-5.16 vs. 74.53+/-4.93 microm). VIP immunogenicity in cells (ganglion and chromaffin) and fibers was robust in fetal adrenal medulla of sheep while nonexistent in baboons. VIP fibers in fetal sheep medulla were smaller in diameter compared to fetal sheep cortex (1.22+/-0.13 vs. 2.93+/-0.34 microm, P<0.05), but not compared to extrinsic nerve fibers (1.30+/-0.09 microm). We hypothesize that in fetal sheep of this age, medullary neurofibers derive primarily from extrinsic sources while cortical fibers arise from cortical ganglion cells. We conclude that at 85% of gestation the potential for VIP neural control of paracrine (e.g., glucocorticoid/catecholamine) interactions in both adrenal cortex and medulla is much greater in fetal sheep compared to fetal baboons.

Adrenal Cortex↗

PubMed1999

Periventricular preoptic area neurons coactivated with luteinizing hormone (LH)-releasing hormone (LHRH) neurons at the time of the LH surge are LHRH afferents.

Earlier studies demonstrated coactivation of the periventricular preoptic area (pePOA) with LHRH neurons at the time of an induced or spontaneous LH surge, suggesting that the pePOA might regulate LHRH neurons. To investigate this hypothesis, studies were conducted to determine the temporal pattern of pePOA Fos activation during the rat estrous cycle and establish the connections of the pePOA neurons with LHRH neurons. Fos activation within LHRH and pePOA neurons showed the same temporal pattern. Both were absent during diestrous I, diestrous II, and the morning of proestrus. Fos was induced in the pePOA and LHRH neurons beginning on the afternoon of proestrus (4 h before lights off), with a decline 8 h later on proestrous evening. Tract-tracing studies then established the relationship between LHRH and pePOA neurons. Retrograde labeling with fluorogold determined that a portion of the Fos-positive pePOA neurons present at the time of the LH surge sent a projection to regions that contain LHRH cells. Anterograde tracer (neurobiotin) injections established that the pePOA neurons sent axons to the LHRH cells. Taken together, these data indicate that the pePOA provides direct input to LHRH neurons that is likely to stimulate LHRH neurons at the time of the LH surge.

Animals↗

PubMed1998

Granulated metrial gland cells contain nitric oxide synthases during pregnancy in the rat.

Nitric oxide synthase (NOS) enzyme activity is present in rat decidua and metrial gland, and the activity decreases near the end of pregnancy. In this study, inducible and endothelial NOS isoforms were immunolocalized to rat granulated metrial gland (GMG) cells using anti-NOS antibodies proven to be isoform specific. These NOS-positive GMG cells are of the natural killer cell lineage as they stained positively for NKR-P1 cell surface receptor, and for perforin. The number of NOS-positive GMG cells corresponded with the level of decidual and metrial gland NOS enzyme activity. NOS activity declined when GMG cells containing NOS decreased in number. Uterine arteriolar vascular smooth muscle also stained positively for inducible NOS and the staining did not change with advancing gestation. Only a minority of uterine myocytes stained positively for inducible NOS and these were subjacent to the placental attachment site. Neuronal NOS immunostaining was not present in the decidua and the metrial gland.

Animals↗

PubMed1997

Progesterone blockade of a luteinizing hormone surge blocks luteinizing hormone-releasing hormone Fos activation and activation of its preoptic area afferents.

Progesterone is capable of facilitating or blocking the luteinizing hormone (LH) surge, depending on the timing of its administration. However, the precise targets of progesterone's actions are unknown. Since recent studies described the presence of a periventricular preoptic area (pePOA) neuron population afferent to LH-releasing hormone (LHRH) neurons that is co-activated to express c-Fos with LHRH neurons at the time of the LH surge, the present study was designed to determine if the pePOA neurons contain progesterone receptors (PRs) and whether progesterone inhibition is manifested by a failure of LHRH and pePOA neurons to become activated at the time of an LH surge. For progesterone facilitation, a group of immature rats each received a silastic capsule (1.57 mm i.d., 3.18 mm o.d., 1.5 cm long) containing estradiol-17beta (E2) in peanut oil (150 microg/ml) at 09.00 h on postnatal day 28 followed 24 h later by a progesterone implant (crystalline, 1.57 mm i.d., 3.18 mm o.d., 1.5 cm long). For progesterone inhibition, a second group of rats received the estrogen capsule and a progesterone capsule (3.35 mm i.d., 4.65 mm o.d., 3.0 cm long) together at 09.00 h on day 28, and 24 h later received only a blank capsule. On the afternoon of postnatal day 29, all animals were anesthetized and perfused for localization of c-Fos and LHRH, PRs alone, or c-Fos and PRs. The present studies determined that following a progesterone-inhibition paradigm, along with blockade of the LH surge, both activation of LHRH and pePOA neurons was low or absent. Staining of PRs in progesterone-facilitated and progesterone-inhibited rats indicated that the pePOA neurons contained PRs in similar patterns. Double labeling of c-Fos and PRs in progesterone-facilitated rats indicated that nearly all the c-Fos-positive neurons of the pePOA (80 +/- 4.2%) co-expressed PRs; in progesterone-inhibited rats, only 32 +/- 12% of few c-Fos-positive neurons also contained PRs. In no instance were LHRH neurons found to contain PRs. Taken together, these data suggest that both progesterone facilitation and inhibition likely involve direct actions of progesterone on the pePOA neurons, and are consistent with a role for the pePOA neurons in transducing steroid effects on LHRH neurons.

Animals↗

PubMed1997

Alpha1-adrenergic receptor blockade blocks LH secretion but not LHRH cFos activation.

Long-standing pharmacological evidence supports a role of alpha-noradrenergic receptors in regulating LH release, yet little is known of the action of these receptors on LHRH neurons at the cellular level. We conducted a series of studies aimed at examining the effects of alpha-adrenergic receptor blockade on LH secretion and the cellular activation of LHRH neurons on proestrus. Our initial study used an irreversible alpha-receptor blocker, phenoxybenzamine (alpha1 > alpha2). A group of proestrous rats were treated with phenoxybenzamine at doses of 20 mg/kg, intraperitoneally, or 2, 10 and 20 mg/kg, intravenously, and compared with vehicle injected controls. Phenoxybenzamine administered intraperitoneally did not completely block the LH surge in all animals, whereas all intravenous doses consistently blocked the LH surge. cFos activation of LHRH neurons, on the other hand, was either unaffected or only slightly reduced by phenoxybenzamine treatment intraperitoneally. The effects of intravenous phenoxybenzamine were different, in that at all dose levels phenoxybenzamine completely blocked the LH surge and reduced by approximately half, the cFos activation in LHRH neurons (independent of dose). The effects of intravenous phenoxybenzamine could be mimicked by substitution of prazosin (an alpha1 antagonist, 4 mg/kg), but not idazoxan (an alpha2 antagonist, 1 mg/kg), administered intravenously at 11.00 h and 13.00 h on proestrus. These data provide evidence that noradrenergic systems operating through alpha1-receptors in the neuronal chain leading to the LH surge, while critical for the release of LHRH at the time of an LH surge, are not responsible for the cFos transcriptional changes in LHRH neurons that accompany the natural LH surge.

Adrenergic alpha-1 Receptor Antagonists↗

PubMed1997

Gonadotropin-releasing hormone neurons: their structure and function.

The gonadotropin-releasing hormone (GnRH) neurons are essential for reproduction. GnRH neurons must release GnRH in a pulsatile pattern for optimal secretion of LH and FSH. In some species, GnRH surges actively evoke the preovulatory LH surge, while in others, GnRH secretion may be permissive. The GnRH neurons have a unique anatomy. The neurons develop, not from the ventricular epithelium like other CNS neurons, but migrate from the olfactory epithelium during fetal life. This feature provides a basis for the clinical link between olfactory and reproductive function and helps explain the atypical distribution of cells and variations in GnRH cell location noted across species. The diffuse organization of GnRH neurons has made the task of understanding the control of GnRH neurons by other CNS neurons more difficult. Recent evidence has helped discriminate influences that are permissive for GnRH release from those that direct changes in GnRH release.

Animals↗

PubMed1995

Activation of the baboon fetal pituitary-adrenocortical axis at midgestation by estrogen: responsivity of the fetal adrenal gland to adrenocorticotropic hormone in vitro.

We have previously demonstrated that increased expression of fetal pituitary proopiomelanocortin mRNA and the induction of enzymes catalyzing fetal adrenal cortisol formation at term are regulated by estrogen-induced changes in placental oxidation of maternal cortisol to cortisone. To test the hypothesis that induction of fetal pituitary adrenocorticotropic hormone (ACTH) production by estrogen-induced changes in placental cortisol oxidation results in increased responsivity of the fetal adrenal gland to ACTH, in the present study we compared fetal adrenal sensitivity to ACTH in vitro at midgestation in untreated controls and in animals treated at this time in gestation with estrogen. Fetal adrenals were obtained on Day 100 (n = 7) and Day 165 (n = 5; term = Day 184) from untreated baboons and on Day 100 following maternal treatment with estradiol (s.c.; Days 70-100; n = 10) or androgen precursor (n = 3). Adrenal slices (15-25 mg) were perifused (100 microliters/min; 37 degrees C) with Medium 199 (no phenol red); media were collected at 10-min intervals and assayed for cortisol and dehydroepiandrosterone. Secretion of cortisol and dehydroepiandrosterone reached equilibrium after 140 min of perifusion; therefore, basal release was calculated as the mean steroid concentrations during 190-240 min. Adrenal slices were then perifused for 20 min with saline or ACTH at 240 (0.001 nmol), 370 (0.01 nmol), and 490 (0.1 nmol) min, and an overall average cortisol/dehydroepiandrosterone secretion rate (pg/min/mg) between 240-600 min was calculated.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenal Cortex↗

PubMed1995

Induction of glucocorticoid receptor expression in hypothalamic magnocellular vasopressin neurons during chronic hypoosmolality.

The site of action for the modulation of hypothalamo-neurohypophyseal function by adrenal steroids is not known. Glucocorticoid receptors (GR) are an obvious potential site of action, but there have been conflicting reports of GR localization in magnocellular neurons. We confirm studies finding undetectable levels of GR in vasopressin or oxytocin neurons, but now report that chronic hypoosmolality induces the expression of GR in magnocellular vasopressin neurons, but not oxytocin neurons. These data support the hypothesis that the vasopressin gene can be directly inhibited by glucocorticoids, and that the induction of GR expression in magnocellular neurons may be part of a redundant set of mechanisms to suppress the expression of AVP during periods of prolonged hypoosmolality. This mechanism represents a novel form of steroid feedback control in brain.

Animals↗

PubMed1994

cFos immunoreactivity is enhanced with biotin amplification.

Through modification of the protocol by Adams, we developed a biotin amplification procedure for immunofluorescence staining of immediate early gene proteins and also applied biotin amplification for metal enhancement of diaminobenzidine staining in an immunoperoxidase protocol. Commercially available anti-cFos antisera were used to compare conventional "Elite" avidin-biotin complex reactions with biotin amplification reactions (visualized with peroxidase staining or streptavidin-Texas Red fluorescence). Biotin amplification and peroxidase staining (with or without nickel salts) enabled detection of cFos in stimulated neurons with primary antibody concentrations five- to tenfold lower than the conventional procedure. With immunofluorescence staining, at primary antibody concentrations too low to detect cFos with the conventional biotin-streptavidin fluorescence staining protocol, biotin amplification enabled clear cFos fluorescence staining with both antisera. The fluorescence staining exhibited a high signal-to-noise ratio and enabled antibody concentrations four times lower than those used for conventional ABC "Elite" peroxidase procedures. In conclusion, the application of biotin amplification to cFos immunocytochemical localization has the promise of aiding the scientist in detecting these immediate early gene products.

3,3'-Diaminobenzidine↗

PubMed1993

Developmental regulation of protein kinase-A and -C activities in the baboon fetal adrenal.

We have previously demonstrated that the estrogen-regulated change in transuteroplacental metabolism of cortisol (F) and cortisone (E) from preferential reduction (E to F) at midgestation to oxidation (F to E) near term results in activation of the hypothalamic-pituitary-adrenal axis of the baboon and the ontogenesis of rate-limiting steroidogenic enzymes, culminating in de novo F secretion. It is well established that transcription of messages activated by peptide-mediated binding to membrane receptors can occur via cAMP-dependent protein kinase-A (PKA) and/or phospholipid/calcium-dependent protein kinase-C (PKC). The present study was designed to determine whether basal levels of PKA and PKC in the fetal adrenal are developmentally regulated during baboon gestation and, thus, could provide the mechanism(s) by which activation of the fetal adrenal near term is mediated. Fetal adrenal glands were obtained on day 100 (n = 8) and day 165 (n = 6) of gestation (term = day 184) from untreated baboons and on day 100 after treatment of the mother with estradiol benzoate, injected sc between days 70-100 to increase estrogen production. PKA activity (picomoles of 32P incorporated into kemptide per min/mg protein) was determined by incubation of adrenal cytosol (12,000 x g; 0.3-30 micrograms protein) in reaction mixtures containing 0.25 mM ATP, 1 x 10(6) dpm [lambda-32P]ATP, and 3 micrograms kemptide in the presence or absence of 0.02 mM cAMP. PKC activity (picomoles of 32P incorporated into histone IIIS per min/mg protein) was determined in cytosol (105,000 x g) and detergent-solubilized membrane fractions after incubation with 0.02 mM ATP, 50 micrograms histone IIIS, and 1 x 10(6) dpm [lambda-32P]ATP in the presence or absence of calcium and phospholipids. Mean (+/- SE) maternal serum estradiol concentrations (nanograms per ml) were 3-fold greater (P < 0.05) at term (1.9 +/- 0.3) than at midgestation and increased (P < 0.05) after treatment with estradiol. PKA activity was greater at term (3965 +/- 546) than at midgestation (2130 +/- 240) and increased (P < 0.05) 2-fold after treatment with estrogen (3525 +/- 416) at midgestation. PKC activity was always 3- to 4-fold lower than that of PKA and was similar in the cytosol and membrane fractions of the cell. In contrast to PKA, cytosolic PKC activity was similar at mid (265 +/- 98)-and late (353 +/- 99) gestation and was not altered by treatment with estradiol (223 +/- 27).(ABSTRACT TRUNCATED AT 400 WORDS)

Adrenal Glands↗

PubMed1991

Responsivity of the baboon fetal pituitary to corticotropin-releasing hormone in utero at midgestation.

The present study was designed to determine whether the baboon fetal pituitary at midgestation was responsive in utero to a bolus injection of CRH. On day 100 of gestation (term = day 184), baboons were anesthetized with halothane/nitrous oxide, the fetus was exteriorized, and a cannula was inserted into a fetal carotid artery. Five minutes later (experimental time zero), a fetal carotid blood sample was obtained, and saline (0.5 ml) with (n = 6) or without (n = 3) ovine CRH (100 ng estimated to equal 500 ng/kg BW) was then infused via the fetal carotid over a 3-min period. Fetal blood samples were taken 5, 15, 30, 45, and 60 min post-CRH/saline treatment and assayed for ACTH. Mean (+/- SE) pretreatment fetal plasma ACTH concentrations were similar in animals that subsequently received saline (26 +/- 3 pg/ml) or CRH (29 +/- 6 pg/ml). Fetal plasma ACTH remained constant after the infusion of saline. In contrast, CRH increased (P less than 0.05) fetal plasma ACTH within 5 min in six of six baboons to a value (58 +/- 12 pg/ml) that exceeded (P less than 0.05) the zero time value and the respective mean value (27 +/- 5 pg/ml) in saline-treated fetuses. Fetal plasma ACTH concentrations continued to rise in four of six baboons 15 min after CRH injection to a level (68 +/- 15 pg/ml) which exceeded that in saline controls (27 +/- 2 pg/ml). In fetuses treated with CRH, overall mean fetal plasma ACTH concentrations from 0-60 min increased at a rate (1.47 pg/min) greater (P less than 0.05) than that in fetuses injected with saline (0.07 pg/min). In contrast to the effects of intracarotid CRH injection, fetal plasma ACTH was not increased after the infusion of 100 ng CRH into a fetal antecubital vein of three additional animals. Collectively, these findings indicate that intracarotid injection of a bolus of CRH into the baboon fetus rapidly increased fetal plasma ACTH concentrations. Moreover, the site of action of CRH was presumably the fetal pituitary. Therefore, we suggest that the baboon fetal hypothalamic-pituitary axis at midgestation has the capacity to secrete ACTH in response to a challenge of CRH.

Adrenocorticotropic Hormone↗

PubMed1990

Concentrations of 13, 14-dihydro-15-keto prostaglandin F2 alpha after pulsatile progesterone administration at the time of luteolysis of heifers.

Progesterone was administered in pulses to 12 dairy heifers from days 17.5 to 22.5 post-estrus in order to determine its ability to modify secretion of PGF2 alpha around the time of luteolysis. Control heifers exhibited pulses of PGFM concomitant with a sharp decline in progesterone concentrations and thus these pulses were temporally associated with luteolysis. Additional pulses of PGFM were observed in heifers receiving exogenous progesterone, but these were not statistically predictable by either dose of progesterone (50 or 100 micrograms) or time of administration (3 or 6 hour intervals). However, all heifers (4/4) treated with progesterone at 3 hour intervals had additional pulses of PGFM as compared to only one heifer (1/4) treated at 6 hour intervals. When pulses of PGFM were induced by exogenous progesterone there was a substantial lag time between the initiation of progesterone treatment and their occurrence. The limited response to progesterone administration and the lack of synchrony is not consistent with an ability of exogenous progesterone to directly stimulate secretion of PGF2 alpha at the time of luteolysis.

Animals↗