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P H Andersen

Publications and source records attributed to P H Andersen.

At least 37 records · Page 2Linked to original sources
PubMed1998

Ipamorelin, the first selective growth hormone secretagogue.

The development and pharmacology of a new potent growth hormone (GH) secretagogue, ipamorelin, is described. Ipamorelin is a pentapeptide (Aib-His-D-2-Nal-D-Phe-Lys-NH2), which displays high GH releasing potency and efficacy in vitro and in vivo. As an outcome of a major chemistry programme, ipamorelin was identified within a series of compounds lacking the central dipeptide Ala-Trp of growth hormone-releasing peptide (GHRP)-1. In vitro, ipamorelin released GH from primary rat pituitary cells with a potency and efficacy similar to GHRP-6 (ECs) = 1.3+/-0.4nmol/l and Emax = 85+/-5% vs 2.2+/-0.3nmol/l and 100%). A pharmacological profiling using GHRP and growth hormone-releasing hormone (GHRH) antagonists clearly demonstrated that ipamorelin, like GHRP-6, stimulates GH release via a GHRP-like receptor. In pentobarbital anaesthetised rats, ipamorelin released GH with a potency and efficacy comparable to GHRP-6 (ED50 = 80+/-42nmol/kg and Emax = 1545+/-250ng GH/ml vs 115+/-36nmol/kg and 1167+/-120ng GH/ml). In conscious swine, ipamorelin released GH with an ED50 = 2.3+/-0.03 nmol/kg and an Emax = 65+/-0.2 ng GH/ml plasma. Again, this was very similar to GHRP-6 (ED50 = 3.9+/-1.4 nmol/kg and Emax = 74+/-7ng GH/ml plasma). GHRP-2 displayed higher potency but lower efficacy (ED50 = 0.6 nmol/kg and Emax = 56+/-6 ng GH/ml plasma). The specificity for GH release was studied in swine. None of the GH secretagogues tested affected FSH, LH, PRL or TSH plasma levels. Administration of both GHRP-6 and GHRP-2 resulted in increased plasma levels of ACTH and cortisol. Very surprisingly, ipamorelin did not release ACTH or cortisol in levels significantly different from those observed following GHRH stimulation. This lack of effect on ACTH and cortisol plasma levels was evident even at doses more than 200-fold higher than the ED50 for GH release. In conclusion, ipamorelin is the first GHRP-receptor agonist with a selectivity for GH release similar to that displayed by GHRH. The specificity of ipamorelin makes this compound a very interesting candidate for future clinical development.

Adrenocorticotropic Hormone↗
PubMed1998

Rumen motility during induced hyper- and hypocalcaemia.

Rumen motility was recorded on an experimental cow by means of telemetric signal transfer from strain gauge force transducers fixed surgically on the peritoneal surface of the rumen wall in the left flank. The normocalcaemic cow was given a standard milk fever treatment with calcium borogluconate (400 ml with 14 mg Ca/ml) intravenously. Transient clinical signs were: decreased rumination, muscle ticks, salivation and a heart rate reduction of 20%. Rectal temperature remained unaltered. Frequency of rumen contractions was reduced up to 40% whereas amplitude of contractions did not deviate from baseline values. Hypocalcaemia was induced in a second experiment by iv infusion of Na2EDTA. At 0.60 mmol/l ionized blood calcium periods of no motility were recorded whereas inactivity of rumen activity was persistent at 0.55 mmol/l ionized blood calcium. The cow went down at 0.45-0.48 mmol/l ionized blood calcium at which point the heart rate was increased by 40%. The high sensitivity of the method employed allowed the conclusion that already at a concentration of ionized blood calcium at 1.0 mmol/l both frequency and amplitude of rumen contractions decreased rapidly although eating behaviour and rumination appeared unaffected during the short term observation periods. Implications of this finding towards health and production in transition cows are discussed.

Animals↗
PubMed1997

Surface expression of the AMPA receptor subunits GluR1, GluR2, and GluR4 in stably transfected baby hamster kidney cells.

The surface expression of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-type glutamate receptor (GluR) subunits GluR1, GluR2, and GluR4 was studied in cultures of stably transfected baby hamster kidney (BHK)-570 cells. Two methods were used to quantify surface expression: cross-linking with the membrane-impermeant reagent bis (sulfosuccinimidyl) suberate (BS3) and labeling of surface receptors with the membrane-impermeant biotinylating reagent sulfosuccinimidyl 2-(biotinamido)ethyl-1,3-dithiopropionate (NHS-ss-biotin) followed by precipitation with neutravidin beads. Western blot analyses of control versus treated cultures revealed that, for all three GluR subunits examined, 25-40% of the total GluR population is located in the plasma membrane of the BHK-570 cells. This finding was corroborated by analyses of the surface expression of [3H]AMPA binding sites in the GluR-expressing BHK-570 cells performed via the biotinylation/precipitation method; these studies revealed that 30-40% of the total binding site population is found in the plasma membrane. Analyses of combinations of the subunits, both GluR1 + GluR2 and GluR2 + GluR4, revealed that heteromeric combinations of the subunits are not trafficked to the surface more efficiently than homomeric receptors. For each of the three subunits, western blots revealed two distinct bands; removal of surface receptors reduced immunoreactivity for the upper band of each subunit by > 90%, whereas immunoreactivity for the lower band was reduced by only 10-20%. Treatment of extracts from the various cell lines with glycopeptidase F resulted in the collapse of the two bands into a single band of lower molecular weight, suggesting that the two original bands represent differentially glycosylated forms of the same polypeptides. These data indicate that the majority of the stably expressed GluR subunits in these cell lines are incompletely glycosylated and that complete glycosylation is associated with trafficking of the GluR subunits to the cell surface.

Animals↗
PubMed1997

Effects of long-term total fasting and insulin on ob gene expression in obese patients.

In the present study the effect of long-term fasting (6 days) on obese (ob) gene expression was examined in nine severely obese females of 34 +/- 3 years and with a body mass index of 46.4 +/- 2.3 kg/m2. Six days of fasting induced a significant weight loss (126.8 +/- 5.3 vs 120.5 +/- 5.1 kg, P < 0.0001). Insulin-stimulated glucose uptake (hyperinsulinemic, euglycemic clamp, insulin infusion rate 1.5 mU/kg per min) was markedly reduced following fasting (M-value 5.96 +/- 0.74 vs 2.79 +/- 0.23 mg/kg per min, P < 0.0001). Ob mRNA/beta-actin concentration in fat biopsies from abdominal subcutaneous adipose tissue was unchanged after 6 days of fasting (1.50 +/- 0.40 vs 1.47 +/- 0.36 arbitrary units, not significant), whereas serum leptin levels decreased significantly from 53.8 +/- 4.7 to 30.7 +/- 2.0 ng/ml (P < 0.0001) during the same period. No significant correlations were found between insulin-stimulated glucose uptake and serum leptin concentration, either prior to the fast or after the fast. Serum leptin levels were unchanged by hyperinsulinemia for 3 h during the clamp prior to the fast, while hyperinsulinemia for 3 h after 6 days of fasting increased serum leptin by 25% (P < 0.01). In conclusion, 6 days of fasting reduced serum leptin by about 40%. In contrast, ob mRNA in abdominal subcutaneous adipose tissue was unchanged. Furthermore, after 6 days of fasting insulin was able to increase the serum level of leptin significantly, indicating that the effect of insulin on the level of leptin is dependent on the nutritional state.

Adipose Tissue↗
PubMed1996

Stable expression of homomeric AMPA-selective glutamate receptors in BHK cells.

cDNAs encoding glutamate receptor glu1, glu2 (Q and R) or glu4 under control of a constitutively active metallothionine promoter, were transfected into baby hamster kidney cells. Following the addition of selection agent, transfectants expressing high levels of glutamate receptor as measured by [3H]alpha-amino-3-hydroxyl-5-methyl-isoxalazole-4-propionate (AMPA) binding, were selected for further studies. Using glutamate receptor antibodies, the receptor proteins were visualized in Western blotting as having a molecular weight of approximately 100 kDa. [3H]AMPA binding to the glutamate receptor expressing cell lines revealed that glu1, glu2 (Q), and glu4 receptors displayed a single site in Scatchard analysis with Kd values of 12, 15.7 and 21 nM, respectively. However, the Ca2+ impermeable variant of the glu2 receptor, glu2 (R) displayed a curvilinear Scatchard plot. Computer resolution suggested the presence of a high and low affinity state (KH = 2.9 nM; KL = 40.7 nM). The pharmacological profile of the [3H]AMPA binding to these recombinant receptors resembled the high affinity [3H]AMPA binding site in rat brain showing high affinity for glutamate, quisqualate, and medium affinity for 6-cyano-7-nitro-quinoxaline-2,3-dione, CNQX; 6,7-dinitro-quinoxaline-2,3-dione, DNQX; and 6-nitro-7-sulphanyl-benzo(f)quinoxaline-2,3,dione, NBQX. Kainate displayed low affinity and N-methyl-D-aspartate (NMDA), was inactive in inhibiting specific [3H]AMPA binding. These cell lines will prove to be important tools in the study of glutamate receptors.

Animals↗
PubMed1996

Anticonvulsant activity of novel derivatives of 2- and 3-piperidinecarboxylic acid in mice and rats.

The relative ability of derivatives of 2-piperidinecarboxylic acid (2-PC; pipecolic acid) and 3-piperidinecarboxylic acid (3-PC; nipecotic acid) to block maximal electroshock (MES)-induced seizures, elevate the threshold for electroshock-induced seizures and be neurotoxic in mice was investigated. Protective index (PI) values, based on the MES test and rotorod performance, ranged from 1.3 to 4.5 for 2-PC benzylamides and from < 1 to > 7.2 for 3-PC derivatives. PI values based on elevation of threshold for electroshock-induced seizures and rotorod performance ranged from > 1.6 to > 20 for both types of derivatives. Since preliminary data indicated that benzylamide derivatives of 2-PC displace [3H]1-[1-(2-thienyl)-cyclohexyl]piperidine (TCP) binding to the phencyclidine (PCP) site of the N-methyl-D-aspartate (NMDA) receptor in the micromolar range and such low affinity uncompetitive antagonists of the NMDA receptor-associated ionophore have been shown to be effective anticonvulsants with low neurological toxicity, the 2-PC derivatives were evaluated in rat brain homogenates for binding affinity to the PCP site. Although all compounds inhibited [3H]TCP binding, a clear correlation between pharmacological activity and binding affinity was not apparent. Select compounds demonstrated minimal ability to protect against pentylenetetrazol-, 4-aminopyridine- and NMDA-induced seizures in mice. Corneal and amygdala kindled rats exhibited different sensitivities to both valproic acid and the nonsubstituted 2-PC benzylamide, suggesting a difference in these two models. Enantiomers of the alpha-methyl substituted benzylamide of 2-PC showed some ability to reduce seizure severity in amygdala kindled rats.

4-Aminopyridine↗
PubMed1996

Identification of oestrogen receptors and oestrogen receptor mRNA in human adipose tissue.

The distribution of adipose tissue has a major impact on the morbidity and mortality associated with obesity. Furthermore, the distribution of adipose tissue seems to be regulated by sex hormones. Controversy exists over whether the effects of sex hormones (oestrogen and testosterone) on human adipose tissue are an indirect or a direct effect as contradictory results have been obtained when investigating the existence of these receptors in human adipose tissue. In the present study the authors reinvestigated the possible existence of oestrogen receptors (ERs) in human adipose tissue. Human adipocytes from both genders were found to contain specific oestrogen binding sites determined by ligand-binding techniques. The binding protein had a molecular weight of 65 kD (which is similar to that of the ER found elsewhere) and it was found that adipocytes contained mRNA encoding the ER. Moreover, human preadipocytes had no oestrogen-binding capacity and did not possess mRNA encoding the ER. Finally, the authors detected regional differences in receptor density. Women had an equal oestrogen-binding capacity in adipose tissue from the subcutaneous abdominal and the visceral depot, whereas men had twice as high oestrogen-binding capacity in subcutaneous adipose tissue compared with adipose tissue in the visceral fat depot. These findings indicate that mature human adipocytes possess ERs and thus, might be an oestrogen-responsive tissue and that oestrogen may be acting directly in mature adipocytes via its specific receptor. Human preadipocytes, however, seemed not to be an oestrogen-responsive tissue. Finally, preliminary data suggest that there might be differences in ER densities in different fat depots.

Abdomen↗
PubMed1996

Studies on in vivo endotoxin plasma disappearance times in cattle.

Endotoxin plasma disappearance (EPDT) times were determined by a modified Limulus amoebocyte lysate (LAL) assay technique after the intravenous administration of 25 micrograms E. coli 055:B5 endotoxin per kg b.w. to 22 Jersey cows. Clinically healthy cows (n = 6) cleared endotoxin from the plasma within 30 min. Cows pretreated with flunixin meglumine (n = 6) had 2-3 times longer plasma disappearance times, while cows pretreated with phenylbutazone (n = 6) had plasma disappearance times which were 6-12 times longer than the healthy control group. A fourth group comprised clinical cases of spontaneously developed hepatic lipidosis (n = 4). None of these cows were able to clear the injected endotoxin dose and one died before the end of the experiment. The acute phase response, described by leukocyte and thrombocyte counts and plasma glucose and zinc concentrations, was not statistically different between the four groups.

Animals↗
PubMed1996

Prevalence of antibodies to lipid A in Danish cattle.

A cross-sectional study was performed on the occurrence of IgG antibodies to lipid A of the Gram-negative bacterial lipopolysaccharide (LPS, endotoxin) on serum of 2272 cattle distributed on 19 Danish dairy herds. The relationship between the concentration of antibodies to lipid A (ALI) and age, herd, pregnancy rate and occurrence of mastitis, bovine virus diarrhoea (BVD), reproductive and digestive disorders, diarrhoea, pneumonia, foot disorders, various infections and traumatic udder lesions was investigated. ALI generally was low in calves and increased during their first 1.5 years of life to a steady state, which could be altered by the occurrence of disease. There were significant differences in the mean ALI among the herds (P < 0.001). High ALI was associated with a low herd pregnancy rate, to preceding occurrence of mastitis (P < 0.048), BVD (P < 0.01), reproduction diseases (P < 0.041) and digestion disorders (P < 0.064) in animals older than 2 years. The calf mortality rate was not associated to ALI and there was no correlation between the ALI in calves and their dams. The occurrence of high ALI levels on a herd basis may be an indication of increased challenge or enhanced immunological defense to Gram-negative bacteria or endotoxin.

Aging↗
PubMed1996

High dissociation constants (pKa) of basic permeants are associated with in vivo skin irritation in man.

Previously, we have investigated the relationship between dissociation constant (pKa) and skin irritation potential. In the present experiment, 12 basic compounds, with varying pKa values ranging from 1.4 to 11.2, were applied on the backs of 12 healthy adult panellists. Cutaneous reactions were measured objectively, using reflectance spectroscopy and transepidermal water loss (TEWL), and evaluated subjectively with a modified Draize scale. A positive correlation between increasing pKa and skin irritation capacity, measured either visually or by reflectance spectroscopy, was found, but only mecamylamine (pKa = 11.2) induced a significant increase in transepidermal water loss. Compounds with low pKa also induced a paradoxical vasoconstriction measured by reflectance spectroscopy. Only high pKa appeared predictive of in vivo skin irritation, and these chemicals apparently induce skin irritation with only minimal disruption of the skin barrier. A simple 1-variable model is predictive of skin irritation for this series of organic permeants with increasing pKa.

Adult↗
PubMed1995

The importance of two specific domains in ligand binding to the AMPA/kainate glutamate receptors GluR2 and GluR6.

Chimeric receptor subunits of the AMPA receptor subunit GluR2 and the kainate receptor subunit GluR6 were constructed and stably expressed in baby hamster kidney cells. By using Ca2+ imaging and radioligand binding, we demonstrated that substitution of a specific domain showing homology to a bacterial leucine-isoleucine-valine binding protein (LIVBP) had no effect on the affinities of the tested agonists, but decreased the affinities of the antagonists CNQX, DNQX, and NBQX. On the other hand, when the first of two domains showing homology to a bacterial glutamine binding protein (QBP) in GluR2 was substituted with the corresponding region from GluR6, the affinity of AMPA decreased sevenfold and the affinity of kainate increased fourfold, indicating the importance of this domain in binding of these agonists. In contrast to this, the affinities of quisqualate and domoate, two other agonists, were unchanged, indicating that a region located C-terminal to the QBP domain is also involved in agonist binding.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMed1995

Functional importance of the carboxyl tail cysteine residues in the human D1 dopamine receptor.

To assess the importance of the cysteine residues Cys347 and Cys351 in the carboxylic tail in the human D1 dopamine receptor, seven mutant receptors were constructed by PCR. The pharmacological and functional properties of the wild-type and mutant receptors were assessed following transient expression in COS-7 cells. Affinities for [3H]SCH 23390 of mutant S347 (Cys347-->Gly), T348 (Tyr348-->stop), S351 (Cys351-->Gly), T351 (Cys351-->stop), T352 (Pro352-->stop), and S347/S351 (Cys347-->Gly and Cys351-->Gly) were similar to that of wild-type receptor, whereas the expression levels were reduced up to 80%. The potency of dopaminergic antagonists for these mutant receptors was very similar to that of the wild-type receptor. However, mutant T347 (Cys347-->stop) showed a 15-25-fold reduced affinity for the antagonists SCH 23390, (+)-butaclamol, and cis-flupentixol, thus not allowing radioligand analysis. Wild-type and mutant receptors responded dose-dependently with similar potency to dopamine and SKF 38393 with an increased adenylyl cyclase activity. However, mutant receptors with the Cys347 residue changed or removed displayed a diminished ability to activate adenylyl cyclase. Dopamine preexposure desensitized wild-type and mutant S351 receptors. However, mutant receptors with Cys347 replaced or the distal part of the carboxyl tail removed were unable to desensitize. Thus, Cys347 in the cytoplasmic tail of the human D1 dopamine receptor is important for the receptor in maintaining the conformation for antagonist binding, to play a crucial role in activation of adenylyl cyclase, and to be essential for agonist-induced desensitization.

Adenylyl Cyclases↗
PubMed1995

Skin irritation in man: a comparative bioengineering study using improved reflectance spectroscopy.

Variable types of skin irritation were induced in 8 human female volunteers, ranging from subclinical to visible erythema with slight oedema. Skin reactions were graded clinically and objectively using transepidermal water loss (TEWL), laser Doppler flowmetry (LDF) and improved reflectance spectroscopy. This last technique enables separation of in vivo erythemas into relatively deoxygenated (venous--deoxy hem) and oxygenated (arterial--oxy hem) haemoglobin components. Compared to uninvolved skin, an empty patch increased oxy hem by 197% +/- 121% (p < 0.05). Exposure to vehicles also changes skin biophysics. At sodium lauryl sulfate (SLS) and hydrochloric acid (HCl) exposed sites, a linear correlation between concentration and oxy hem, LDF and TEWL was found. These chemicals predominantly increased TEWL values. Nonanoic acid (NON) and imipramine (IMI) also raised oxy hem, LDF and TEWL values linearly at increasing concentrations. Although IMI 2.5% clinically was graded as a type ++ response, no significant increase in TEWL was found. The improved reflectance spectroscopic technique proved valid in skin irritation studies, with a higher sensitivity than laser Doppler flowmetry, and allowed irritant vascular reactions to be discriminated into arterial and venous components. Furthermore, our observations clearly demonstrate that clinically indistinguishable skin irritation reactions induce significantly different changes in barrier function (disruption) and vascular status.

Adult↗
PubMed1994

Stable expression of a functional GluR6 homomeric glutamate receptor channel in mammalian cells.

This study demonstrates the stable expression of a functional ionotropic glutamate receptor in a mammalian cell line of non-neuronal origin. The kainate-selective glutamate receptor GluR6 was constitutively expressed under the control of a metallothionein promoter. Clones were isolated expressing approximately 3 pmol of receptor per mg of protein. Functionality of the recombinant GluR6 was demonstrated both by electrophysiology and by Ca2+ imaging. Application of kainate to the GluR6-transfected cells activated an inward current response at a holding potential of -60 mV. The kainate concentration needed to evoke 50% of the maximal response (EC50) was calculated to be 0.82 +/- 0.39 microM. The current-voltage relationship was found to be almost linear, with a reversal potential of -2.5 +/- 4.8 mV. Application of kainate also resulted in an increase in the intracellular Ca2+ concentration measured by Ca2+ imaging. The pharmacological profile of [3H]kainate binding to the recombinant GluR6 resembled the high-affinity [3H]kainate binding sites in rat brain, showing high affinity for domoate (Ki = 5.1 +/- 3.0 nM) and kainate (Kd = 12.9 +/- 2.4 nM). No decrease in GluR6 expression level was observed over > 75 passages of the transfected cells. When domoate, a slowly desensitizing GluR6 agonist, was included in the growth medium for 3 weeks, the number of GluR6 binding sites decreased by 30%, indicating the importance of complete channel closure for stable expression.

Animals↗
PubMed1994

Characteristics of stably expressed human dopamine D1a and D1b receptors: atypical behavior of the dopamine D1b receptor.

Human dopamine D1a and D1b receptors were stably expressed in Baby Hamster Kidney (BHK) or Chinese Hamster Ovary (CHO) cells. [3H]SCH23390 saturation experiments indicated the presence of only a single binding site in the D1a expressing cell line with a Kd of 0.5 nM. In D1b expressing cell lines, two binding sites were observed with Kd values of 0.5 and 5 nM in CHO cells and 0.05 and 1.6 nM in BHK cells, respectively. Neither of the receptors affected Ca2+ metabolism whereas they both were coupled in a stimulatory fashion to adenylyl cyclase. The pharmacological profile of both the D1a and D1b receptors as assessed from inhibition of specific [3H]SCH 23390 binding was classical D1-like. Thus, benzazepine derivatives as well as the atypical neuroleptics, clozapine and fluperlapine, exhibited high affinity whereas D2 selective compounds like sulpiride and spiperone had low affinity for these receptors. Besides SCH 23390, only NNC 112, fluphenazine and bulbocapnine were able to discriminate between the two states of the D1b receptor. In case of the D1a receptor, the Ki values obtained in binding experiments were very similar to Ki values obtained from inhibition of dopamine stimulated adenylyl cyclase. In the D1b expressing cell line, the Ki values obtained from inhibition of the dopamine stimulated adenylyl cyclase indicated a significantly better correlation with the state of the D1b receptor showing high affinity for antagonists. In agreement with observations from binding experiments, dopamine was around 20 fold more potent in stimulating adenylyl cyclase via the D1b receptor as compared to the D1a receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl Cyclases↗
PubMed1994

Expression of glycogen synthase and phosphofructokinase in muscle from type 1 (insulin-dependent) diabetic patients before and after intensive insulin treatment.

The aim of the present study was to determine whether short-term appropriate insulinization of Type 1 (insulin-dependent) diabetic patients in long-term poor glycaemic control (HbA1C > 9.5%) was associated with an adaptive regulation of the activity and gene expression of key proteins in muscle glycogen storage and glycolysis: glycogen synthase and phosphofructokinase, respectively. In nine diabetic patients biopsies of quadriceps muscle were taken before and 24-h after intensified insulin therapy and compared to findings in eight control subjects. Subcutaneous injections of rapid acting insulin were given at 3-h intervals to improve glycaemic control in diabetic patients (fasting plasma glucose decreased from 20.8 +/- 0.8 to 8.7 +/- 0.8 mmol/l whereas fasting serum insulin increased from 59 +/- 8 to 173 +/- 3 pmol/l). Before intensified insulin therapy, analysis of muscle biopsies from diabetic patients showed a normal total glycogen synthase activity but a 48% decrease (p = 0.006) in glycogen synthase fractional velocity (0.1 mmol/l glucose 6-phosphate) (FV0.1) and a 45% increase (p = 0.01) in the half-maximal activation constant of glycogen synthase (A0.5). The activity of phosphofructokinase and the specific mRNA and immunoreactive protein levels of both glycogen synthase and phosphofructokinase were similar in the two groups. The 2.8-fold increase in serum insulin levels and the halving of the plasma glucose level for at least 15 h were associated with a normalization of glycogen synthase fractional activity (FV0.1) and of the half-maximal activation constant (A0.5) whereas the enzyme activity of phosphofructokinase and the mRNA and protein levels of both glycogen synthase and phosphofructokinase remained normal.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗