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K A Berg

Publications and source records attributed to K A Berg.

At least 37 records · Page 2Linked to original sources
PubMed1994

Signal transduction differences between 5-hydroxytryptamine type 2A and type 2C receptor systems.

The cDNAs for human 5-hydroxytryptamine (5-HT)2C and 5-HT2A receptors were stably transfected separately into parent Chinese hamster ovary cells, and cell lines in which levels of transfected receptor protein expression and accumulation of inositol phosphates in response to 5-HT were comparable were chosen for study. The effect of activation of these receptors on 5-HT1B-like receptor-mediated responsiveness (i.e., inhibition of forskolin-stimulated cAMP accumulation) was studied. Activation of 5-HT2C receptors with 5-HT (0.1-100 microM) abolished the 5-HT1B-like response, which returned when 5-HT2C receptors were blocked with mesulergine (1 microM). Furthermore, the maximal response to 5-carboxytryptamine was reduced in a concentration-dependent manner by the 5-HT2A/5-HT2C-selective partial agonist (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane. In contrast, activation of 5-HT2A receptors with either 5-HT or (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane did not alter the 5-HT1B-like response. The reduction of 5-HT1B-like responsiveness produced by 5-HT2C receptor activation was independent of protein kinase C activation and increases in the intracellular calcium concentration. Although 5-HT2A and 5-HT2C receptors are strikingly similar in structure and pharmacology, and the signal transduction systems coupled to these receptors have been thought to be similar, if not identical, these data provide the first evidence for fundamental differences in the signal transduction systems of these 5-HT2 receptor subtypes.

Amphetamines↗
PubMed1994

5-Hydroxytryptamine type 2A receptors regulate cyclic AMP accumulation in a neuronal cell line by protein kinase C-dependent and calcium/calmodulin-dependent mechanisms.

The effects of 5-hydroxytryptamine (5-HT)2A receptor activation on cAMP formation were studied in a cell line derived from embryonic rat cortex (A1A1). 5-HT (EC50 = 0.87 microM) amplified the amount of cAMP formed in response to 5'-N-ethylcarboxamidoadenosine (an adenosine A2 receptor agonist), cholera toxin, and forskolin after 15 min of coincubation in the presence of the phosphodiesterase inhibitor rolipram. This effect of 5-HT was blocked by 10 nM ketanserin as well as by 10 nM spiperone, indicating a response mediated by the 5-HT2A receptor subtype. Similarly, cAMP accumulation was enhanced by coincubation with the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA) and the calcium ionophore A23187. After exposure to PMA for 24 hr (PKC-depleted cells), 5-HT and A23187 still enhanced cAMP formed in response to forskolin and 5'-N-ethylcarboxamidoadenosine, whereas the amplifying effects of PMA were abolished. Analysis by Western blots and PKC activity measurements revealed that, of three PKC isoforms detected in A1A1 cells (alpha, delta, and epsilon), only the calcium-independent isoform PKC-epsilon remained in membrane fractions after long term PMA treatment. In PKC-depleted cells, 5-HT-mediated amplification was greatly reduced after treatment with the calcium chelator 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (acetoxymethyl)-ester or the calmodulin antagonists calmidazolium and N-(6-aminohexyl)-5-chloro-1-napthalenesulfonamide hydrochloride. In addition, 5-HT-mediated amplification of cAMP accumulation was reduced by the PKC inhibitor staurosporine in normal cells but was unaffected in PKC-depleted cells. In conclusion, these data suggest that 5-HT2A receptor activation can amplify cAMP formation in A1A1 cells by two distinct pathways coupled to the hydrolysis of inositol phosphates, i.e., PKC and calcium/calmodulin.

Adenosine↗
PubMed1993

Evaluation of spermatological parameters used to predict the fertility of frozen bull semen.

Post-thaw motility, velocity and acrosome integrity of frozen semen were determined in 18 bulls with varying fertility (average non-return rates: 71.3 (+/- 2.8)--range: 65.2-75.7). Five semen straws were investigated from each bull. The average values for sperm motility (percentage motile spermatozoa), sperm velocity (graded from 0-3) and acrosome integrity (proportion of spermatozoa with intact acrosome) were 67.5%, 2.5 and 79.3%, respectively. Significant correlations were found between sperm motility and velocity, but not between sperm motility and acrosome integrity. Both sperm motility and velocity were significantly related to bull fertility. It was concluded that of the post-thaw semen characteristics investigated in this study these 2 parameters provided a reliable basis for prediction of bull fertility.

Acrosome↗
PubMed1993

5-Hydroxytryptamine1-like receptors linked to increases in intracellular calcium concentration and inhibition of cyclic AMP accumulation in cultured vascular smooth muscle cells derived from bovine basilar artery.

Vascular smooth muscle cells derived from bovine basilar artery by the explant method were grown in culture. In the presence of 1 microM forskolin and the phosphodiesterase inhibitor rolipram, 5-hydroxytryptamine (5-HT) agonists inhibited by 90 to 100% the accumulation of intracellular cyclic AMP (cAMP) with a rank order of potency 5-carboxamidotryptamine (5-CT) > or = 5-HT > 5-benzyloxytryptamine = sumatriptan > RU24969 [5-methoxy-3(1,2,3,6-tetrahydro-4-pyridinyl)-1H indole succinate] > (+/-)-8-hydroxydipropylaminotetralin. In suspensions of cells loaded with the calcium-sensitive probe fura-2, 5-CT and 5-HT caused a biphasic increase in the concentration of intracellular free calcium ([Ca++]i) that consisted of both transient and sustained phases. The transient phase was reduced and the sustained phase abolished in the absence of extracellular calcium. The EC50 for 5-CT-induced increase in [Ca++]i (6 nM) was similar to that for inhibition of cAMP accumulation (1.3 nM). Both the inhibition of cAMP accumulation and increase in [Ca++]i were inhibited by the antagonist methiothepin (pA2 = 8.9), but not by the antagonists ketanserin, spiperone and pindolol. Both the inhibition of cAMP accumulation and increase in [Ca++]i were attenuated by greater than 85% in cells that were pretreated with pertussis toxin. PI turnover was not stimulated by 5-CT. The rank order of agonist potency, as well as the antagonist sensitivity, indicates responses mediated by one or more 5-HT1-like-type receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

BrdUrd incorporation studies for evaluation of spermatogenesis in the blue fox.

The utility of BrdUrd incorporation techniques for studies of spermatogenesis was investigated in the blue fox. BrdUrd was injected intraperitoneally followed by collection of testicular tissue by castration/hemicastration at intervals up to 35 days after pulse labelling. Fluorescent tagged monoclonal antibodies against BrdUrd allowed detection of cells with incorporated tracer in histological sections by fluorescent light microscopy as well as in isolated testicular cells by bivariate BrdUrd/DNA flow cytometry. The duration of the spermatogenic cycle was estimated by following the labelled cohort of preleptotene spermatocytes by immunofluorescence in sections through the various stages of maturation to the late spermatid stage. These data were confirmed by bivariate BrdUrd/DNA flow cytometry of testicular cells in suspensions. Furthermore, estimations of the S phase durations and length of the spermatogonial cell cycle were possible. A consistent and satisfactory fluorescence intensity of incorporated label throughout the study shows that degradation of the incorporated label is no practical problem for this type of study, and suggests that the method is an excellent tool for studying aspects of proliferation and maturation during normal as well as perturbed spermatogenesis. Advantages of the described method include avoidance of potential radiation influence on spermatogenesis from commonly used radiolabelled tracers, e.g., 3H-TdR, and that both large and small animals can be investigated at modest cost since the unlabelled BrdUrd is considerably less expensive than labelled tracers.

Animals↗
PubMed1989

Congenital cardiovascular malformations in twins and triplets from a population-based study.

Data from the Baltimore-Washington Infant Study of congenital cardiovascular malformations permitted detailed analysis of congenital cardiovascular malformations in 62 twins and 3 triplets and 2303 singleton cases. A probability sample of controls (n = 2793) included 43 twins. The case prevalence of multiple births was 28 of 1000, compared with a 15 of 1000 prevalence among controls (chi 2 = 5.7). There were more girls among case twins than among case singletons and controls (chi 2 = 9.0). Monozygosity was no more frequent in case twins than in controls. Looping defects occurred in 4 monozygotic twin pairs compared with only 1 dizygotic twin pair. The twinning process itself may be implicated in the development of congenital cardiovascular malformations in some of these infants, especially those with looping defects, but concordance of types of defects in 4 of 65 pairs implicates genetic factors in the determination of some forms of congenital cardiovascular malformations.

Diseases in Twins↗
PubMed1989

Cardiac malformations in relatives of infants with hypoplastic left-heart syndrome.

In a pilot study of relatives of infants with hypoplastic left-heart syndrome (HLHS), we obtained a medical history, cardiovascular examination, and echocardiogram in 48 first-degree relatives of 11 probands with isolated HLHS and 3 with HLHS and noncardiac malformations. Echocardiography confirmed heart defects in 5 of 41 relatives of patients with isolated HLHS. In four instances, the cardiac abnormality was unrecognized. Among 7 relatives of infants with HLHS and extracardiac anomalies, no heart defects were detected. Cardiac defects occurred in first-degree relatives of probands at a frequency higher than previously predicted by an additive multifactorial model of inheritance. These findings suggest that first-degree relatives of HLHS probands may have an increased risk for subclinical cardiac defects and that genetic factors likely contribute to the cause of left-heart blood-flow lesions.

Family↗
PubMed1989

Insulin-like growth factor-I potentiates thyrotropin stimulation of adenylyl cyclase in FRTL-5 cells.

We reported that TSH and insulin-like growth factor-I (IGF-I), which were known to synergistically stimulate DNA synthesis, synergize to elevate the 1,2-diacylglycerol content of FRTL-5 thyroid cells. We presented evidence that cAMP is the proximal mediator of these actions of TSH. To further define the mechanism of this interaction, we investigated the effects of IGF-I on TSH stimulation of adenylyl cyclase. Long and short term effects of IGF-I or high doses of insulin were studied in FRTL-5 cells that were maintained in serum-, hormone-, and growth factor-free medium for 4-7 days (basal cells). When cells were incubated with high doses of insulin for 7 days and acutely stimulated, a 10-fold increase in sensitivity and a 2-fold increase in maximal responsiveness of cAMP accumulation to TSH were observed. To study shorter term effects, cells were preincubated with insulin for 3 h and then exposed to TSH, cholera toxin, or forskolin. Incubation with high doses of insulin for 3 h caused 30-300% increases in cAMP accumulation at high doses of TSH (greater than or equal to 1 mU/ml), cholera toxin (greater than 0.1 microM), and forskolin, but did not affect the EC50 for TSH. Dose-response studies were consistent with insulin acting via receptors for IGF-I, and IGF-I caused a similar effect. There was a 45% increase in adenylyl cyclase activity stimulated by TSH in membranes isolated from cells incubated with high doses of insulin for 3 h. Pretreatment of FRTL-5 cells with pertussis toxin, which ADP-ribosylates the inhibitory G-protein Gi, or adenosine, which we show inhibits cAMP accumulation by interacting with Gi, did not affect insulin/IGF-I enhancement of cAMP accumulation. We suggest that synergism of actions of TSH and IGF-I may in part be due to IGF-I enhancement of TSH stimulation of adenylyl cyclase.

Adenylate Cyclase Toxin↗
PubMed1989

Factors influencing the success rate of artificial insemination with frozen semen in the dog.

The conception rate and fecundity of 36 bitches artificially inseminated with frozen semen from various sources were investigated: 8 bitches were inseminated with semen frozen at the clinic, and 28 were inseminated with imported frozen semen. The mean progressive post-thaw motility of semen was 60%. Fourteen bitches were inseminated once only and 22 bitches were inseminated twice at 1-2-day intervals. An overall conception rate of 67% (24/36) was obtained with a mean litter size of 6.4. One insemination yielded a slightly lower conception rate than 2 inseminations (64 vs 69%). Among bitches which were inseminated twice, non-pregnant bitches had significantly lower mean plasma progesterone concentrations at the second insemination than did pregnant bitches, and this indicates that the second insemination was performed too early. Correct timing of inseminations is therefore essential to obtain pregnancy with frozen semen.

Animals↗
PubMed1988

Combinative ligand-receptor interactions: epinephrine depresses RAW264 macrophage antibody-dependent phagocytosis in the absence and presence of met-enkephalin.

Macrophages have been shown to possess cell surface receptors for opiates and catecholamines. The abilities of these ligands to affect RAW264 macrophage antibody-dependent effector activity directed against sheep red blood cells were tested. Phagocytosis was measured by the uptake of 51Cr labeled erythrocytes and optical microscopy. Cytolysis was measured by 51Cr-release assays. Met-enkephalin increased specific antibody-dependent phagocytosis in a dose-dependent fashion; the optimal dose was found to be 10(-8) M. Epinephrine diminished phagocytosis in a dose-dependent manner exhibiting a maximal inhibition at 10(-4)-10(-5) M. This inhibition can be blocked by propranolol. The combined effects of simultaneous treatment with met-enkephalin and epinephrine were measured. At the several doses tested, the combined effects of these two ligands on the amount of phagocytosis were equivalent to or more inhibitory than epinephrine alone. Thioglycolate-elicited murine peritoneal macrophages demonstrated similar responses to epinephrine, met-enkephalin, and their combination. Therefore, in vitro models more closely approximating in vivo neuroregulation of macrophage function demonstrate phagocytic inhibition.

Animals↗
PubMed1988

Cholesteryl ester handling by RAW264 macrophages: response to native and acetylated low density lipoprotein.

The effects of LDL and Ac-LDL on the growth properties, morphology, and cholesteryl ester (CE) metabolism of the RAW264 macrophage cell line have been characterized. Cells were grown in media supplemented by a defined media (DM) mixture or fetal bovine serum (FBS). The addition of LDL or Ac-LDL to the culture media did not significantly alter cell growth properties. Cytoplasmic deposition of CE was observed by fluorescence microscopy in macrophages treated with LDL or Ac-LDL but not in untreated controls. Dose-response studies have shown that cholesteryl ester (CE) can accumulate in RAW264 treated with LDL. Cellular cholesterol content saturated at 4 hours with 50 micrograms/ml LDL; this effect may be associated with receptor saturation. Dose-response studies conducted with Ac-LDL in DM have shown dramatic increases in total cell cholesterol content. However, deposition of CE was not observed below Ac-LDL concentrations of 100 micrograms/ml. This indicates that a critical concentration of Ac-LDL must be reached to trigger deposition in DM. In contrast, no critical concentration of Ac-LDL was observed in macrophages grown in medium supplemented with 10% FBS. Cholesterol esterification in response to LDL and Ac-LDL was examined by 14C-oleic acid incorporation into CE. These results confirmed the mass cellular cholesterol and CE measurements. Kinetic studies conducted with RAW264 cells treated with 50 or 100 micrograms/ml Ac-LDL resulted in a cholesterol efflux from the cells at 6-12 hours of incubation. Therefore, these studies show that (1) the nature of CE deposition is highly dependent upon the incubation media and (2) CE deposition is very sensitive to Ac-LDL concentration under certain conditions.

Animals↗
PubMed1988

Prenatal detection of cardiovascular malformations by echocardiography: an indication for cytogenetic evaluation.

Prenatal diagnosis of congenital cardiovascular malformations by echocardiography may signal associated chromosome abnormalities. The exact proportion of these associations is not known but is expected to be higher than that with live-birth. To estimate the risk that a fetus with an echocardiographically detected heart defect has an autosomal trisomy or Turner syndrome, we adjusted the known frequency of aneuploidy in live-born infants with congenital cardiovascular malformations by the reported rate of spontaneous abortion, with data from a population-based case-control study of congenital cardiovascular malformations in which 268 cases (12.7%) had both congenital cardiovascular malformations and a chromosome abnormality. Included in the present analysis were 188 aneuploid infants with congenital cardiovascular malformations that would have been detectable by fetal echo. When data are adjusted for the high spontaneous abortion rate of aneuploid fetuses, we estimate that there would have been more than a threefold increase in aneuploidy over the 13% seen at live-birth. Thus cytogenetic analysis is appropriate in a fetus with echo-diagnosed congenital cardiovascular malformations.

Aneuploidy↗
PubMed1988

Thyroid-stimulating hormone and insulin-like growth factor-1 synergize to elevate 1,2-diacylglycerol in rat thyroid cells. Stimulation of DNA synthesis via interaction between lipid and adenylyl cyclase signal transduction systems.

Thyroid-stimulating hormone (TSH) and insulin-like growth factor-1 (IGF-1) synergistically stimulate DNA synthesis in thyroid cells. In this report, a novel mechanism for mediation of this synergistic interaction is described in rat thyroid (FRTL-5) cells. Because phorbol myristate acetate stimulates DNA synthesis, the effects of TSH, IGF-1 and insulin on FRTL-5 cell content of 1,2-diacylglycerol (1,2-DG), the endogenous activator of protein kinase C, were measured. After 6 d, TSH, IGF-1 and insulin caused increases in cellular 1,2-DG (mean +/- SE) to 180 +/- 10%, 540 +/- 50%, and 360 +/- 40% of control, respectively, whereas TSH plus IGF-1 and TSH plus insulin synergistically increased 1,2-DG to 1,890 +/- 310% and 1,690 +/- 230%, respectively. In the absence of insulin, the effect of TSH to elevate 1,2-DG exhibited an EC50 of approximately 2,000 microU/ml. The synergistic interaction of insulin and TSH was found to increase the potency of TSH by 300-fold (EC50 was approximately 7 microU/ml) in addition to increasing the efficacy of TSH. The effect of TSH appeared to be mediated by TSH-stimulated increases in cyclic AMP (cAMP). Forskolin and 8-bromo-cAMP, like TSH, caused modest increases in 1,2-DG and DNA synthesis, whereas forskolin plus insulin and 8-bromo-cAMP plus insulin markedly elevated 1,2-DG content and stimulated DNA synthesis. Under all conditions, increases in 1,2-DG content correlated with stimulation of DNA synthesis. These findings suggest that the synergistic stimulation of DNA synthesis in thyroid cells by TSH, via cAMP, and IGF-1 is mediated by 1,2-DG. Moreover, they implicate a novel interaction between the lipid and adenylyl cyclase signaling systems for the regulation of cell proliferation.

Adenylyl Cyclases↗
PubMed1987

Familial risks of congenital heart defect assessed in a population-based epidemiologic study.

Congenital heart defects (CHD) represent a heterogeneous group of disorders caused by chromosome abnormalities, mendelian disorders, teratogenic exposures, and unknown etiologic mechanisms. A large group of various isolated defects is presumably multifactorial in origin. Previous studies of familial risks for specific anatomic defects obtained from clinical series may include significant biases and obscured pathogenic relationships. In this population-based study we analyzed all cases of CHD in infants and a control birth cohort in the Baltimore-Washington area. The rates of CHD were defined for first-degree relatives of cases with isolated defects, grouped by a pathogenic classification scheme. Precurrence risks were found to vary among the groups, and risks for flow lesions were higher than previously reported. The sibling precurrence risk for hypoplastic left heart syndrome (13.5%) was not significantly different from that expected for an autosomal recessive mechanism; the risks for different types of ventricular septal defects (VSD) varied among mechanistic groups. The results indicate that the additive multifactorial model does not adequately account for the risks in all forms of isolated CHD of unknown etiology.

District of Columbia↗
PubMed1987

A monoclonal antibody (BMA-1) reactive with murine B cells as well as resident and elicited but not activated macrophages.

Hybridoma technology has been employed to prepare a monoclonal antibody that recognizes a subpopulation of mononuclear leukocytes. Enzyme-linked immune assay revealed a cell clone producing a monoclonal antibody reactive with elicited but not activated C57Bi/6 peritoneal macrophages. Detailed analyses using fluorescence flow cytometry demonstrated that this monoclonal antibody binds to B cells, B cell blasts, as well as to the resident and elicited macrophages, but not to activated macrophages, T cells, red blood cells, or syngeneic fibroblasts. This antigen has been designated BMA-1. Antigenic expression is greatest upon resident macrophages. A bimodal level of expression is found on elicited macrophages while activated macrophages possess low levels of expression. The unique cellular distribution of this antigen indicates that it is lost during macrophage differentiation to the activated state. Immunoprecipitation studies indicate that this antigen is composed of multiple subunits; the primary subunit possesses a molecular weight of 38,000. This new tool should be valuable in the analysis of heterogeneous macrophage populations and in defining molecular differentiation pathways.

Animals↗
PubMed1986

Fluorescence studies of macrophage recognition and endocytosis of native and acetylated low-density lipoprotein.

Macrophage recognition and endocytosis of 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (diI)-labeled low-density lipoprotein (LDL) and acetyl LDL (Ac-LDL) was studied using fluorescence flow cytometry and fluorescence video intensification microscopy. RAW264 macrophages and U937 monocytes were grown in the tissue culture media in the presence and absence of LDL and Ac-LDL. Several lines of evidence indicate that receptor-mediated endocytosis of diI-labeled LDL or Ac-LDL was taking place. Binding can be distinguished from binding plus endocytosis by incubation at 4 and 37 degrees C, respectively. Binding was saturable at 4 degrees C and uptake at 37 degrees C was time- and ligand dose-dependent. Also, unlabeled LDL and Ac-LDL compete for their receptors. Macrophages grown in the presence or absence of LDL demonstrated distinct labeling patterns. LDL receptors were significantly increased by culture in defined medium without serum lipoproteins, while Ac-LDL receptors remained unaffected. Flow cytometry can provide an important tool to examine receptor levels, modulation of these levels and receptor-mediated endocytosis. Video intensification microscopy of similarly labeled cells has been performed. Receptors appear as punctate fluorescence, usually distributed randomly across the cell surface.

Acetylation↗
PubMed1986

Seasonal changes in testicular structure and function in the blue fox (Alopex lagopus), as quantified by morphometric analysis and measurement of adenylate cyclase activity.

The volume of the blue fox testis showed 5-fold changes during the year, associated with considerable changes in cellular composition. The seminiferous epithelium was maximally regressed in August, when 94% of tubules contained only spermatogonia. By late October, approximately 6 months before the mating season, 40% of tubules contained primary spermatocytes. From the middle of January until the end of April all tubules contained spermatids or more advanced haploid cells. Tubular diameter increased by 73% during testicular re-development, and epithelial height increased 3-fold. Regression to the basal state occurred during May to July. The volume densities of the seminiferous epithelium and of interstitial tissue remained approximately constant throughout the year. Soluble Mn2+-dependent adenylate cyclase activity showed seasonal variations that paralleled those of the haploid germ cell population and testicular volume, whereas somatic cell adenylate cyclase activity was relatively constant.

Adenylyl Cyclases↗