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J E Wikberg

Publications and source records attributed to J E Wikberg.

At least 91 records · Page 5Linked to original sources
PubMed1996

Characterization of alpha 2-adrenoceptor subtypes in the porcine eye: identification of alpha 2A-adrenoceptors in the choroid, ciliary body and iris, and alpha 2A- and alpha 2C-adrenoceptors in the retina.

The subtypes of alpha 2-adrenoceptor were characterized in the choroid, ciliary body, iris and retina of the pig eye by using radioligand binding. [3H]-MK912 labelled dense populations of alpha 2A-adrenoceptors in the choroid and ciliary body. In the retina, [3H]-MK912 labelled both alpha 2A- and alpha 2C-adrenoceptors. In the iris, receptors of the alpha 2A-adrenoceptor type were detected by using either [3H]-MK912 or [3H]-RX821002 as radioligands.

Animals↗
PubMed1996

Characterization of adrenoceptor types and subtypes in American bullfrogs acclimated to warm or cold temperature.

While indirect evidence suggested that the responsiveness of frog adrenoceptors changes in response to temperature, direct measurement of adrenoceptor binding following acclimation to warm and cold temperatures had not been done. In the present study, the radioligands [3H]prazosin, [3H]RX821002, and [125I]cyanopindolol were used to label and quantify alpha 1-, alpha 2-, and beta-adrenoceptors in bullfrogs acclimated to warm or cold environments. The number of alpha 1-, alpha 2-, and beta-adrenoceptors in atrium, ventricle, and kidney membranes was not significantly different between warm- and cold-acclimated frogs. Characterization of receptor subtypes using pharmacological antagonists demonstrated that alpha 2-adrenoceptors in frog spinal cord and kidney were of the same pharmacological subtype, which is similar to the mammalian alpha 2A-subtype. The beta-adrenoceptor in frog ventricle, atrium, and kidney was the beta 2-subtype. These results suggest that while the alpha 1-, alpha 2-, and beta-adrenoceptor types have evolved in the frog, multiple subtypes of adrenoceptors are not necessary for physiological regulation in this species.

Acclimatization↗
PubMed1996

Localization of ACTH receptor mRNA by in situ hybridization in mouse adrenal gland.

Stimulation of steroidogenesis in the adrenal cortex is a major physiological action of adrenocorticotropic hormone (ACTH). This action is presumed to be mediated by the ACTH receptor and is functionally connected with the hypothalamus-pituitary-adrenal axis. To gain information concerning the distribution of the ACTH receptor in this axis, we examined mRNA for the ACTH receptor in the adrenal gland, pituitary and hypothalamus of the mouse, by using in situ hybridization. The Y-1 mouse adrenal tumour cell line was also examined. The specific hybridization signal for ACTH receptor mRNA was uniformly distributed in the Y-1 cells, although the level of expression was low. The adrenal cortex showed a strong signal for the ACTH receptor mRNA in both the zona fasciculata and the zona glomerulosa, sites that are involved in the mediation of the action of ACTH on the synthesis and release of glucocorticoids and aldosterone. In the zona reticularis of the cortex, a small number of cells showed positive hybridization signals. Moreover, a few scattered cells were positive in the adrenal medulla. In contrast, the hybridization results for both the hypothalamus and the pituitary proved to be negative. These results indicate that the mouse ACTH receptor is a peripheral receptor that is exclusively located in the adrenal gland.

Adrenal Glands↗
PubMed1996

Major pharmacological distinction of the ACTH receptor from other melanocortin receptors.

The mouse adrenocortical cell line Y1, that expresses ACTH receptors (MC2R), was used to probe the binding of ACTH and MSH peptides by using radio-labelled ACTH (1-39). The Y1 cells were found to bind [125I]-labelled ACTH (1-39) with high affinity (Kd approximately 130 pM). However, none of the melanocortin peptides NDP-MSH, alpha-MSH, beta-MSH or gamma 1-MSH could compete with the binding of the labelled ACTH(1-39). When other MC receptor subtype DNAs (MC1, MC3 and MC4) were transfected into the Y1 cells, characteristic binding of the [125I]NDP-MSH appeared for each of the receptor subtype, but no specific binding was present in non-transfected cells. This is the first report clearly demonstrating that the ACTH receptor binds only ACTH, but not other melanocortin peptides.

Adrenal Cortex↗
PubMed1996

Characterisation of the melanocortin 4 receptor by radioligand binding.

The DNA encoding the human melanocortin 4 receptor was expressed in COS (CV-1 origin, SV 40) cells and its radioligand binding properties was tested by using the [125I][Nle4, D-Phe7] alpha-melanocyte stimulating hormone (MSH). The radioligand was found to bind to a single saturable site with a Kd of 3.84 +/- 0.57 nmol/l in the MC4 receptor expressing cells. The order of potency of a number of substance competing for the [125I][Nle4, D-Phe7] alpha-MSH binding was the following; [Nle4, D-Phe7] alpha-MSH > [Nle4]-alpha-MSH > beta-MSH > desacetyl-alpha-MSH > alpha-MSH > ACTH (1-39) > ACTH (4-10) > gamma 1-MSH > gamma 2-MSH. This order of potency is unique for the melanocortin 4 receptor when compared to our previously published data for the other melanocortin receptor subtypes. Most notably the melanocortin 4 receptor shows highest affinity for beta-MSH, among the endogenous MSH-peptides. Furthermore the melanocortin 4 receptor shows very low affinity for the gamma-MSH peptides. This distinguishes the melanocortin 4 receptor from the melanocortin 3 receptor, which is the other major central nervous system melanocortin-receptor, as melanocortin 3 receptor shows high affinity for gamma-MSH. Our finding might indicate a specific role for beta-MSH for the melanocortin 4 receptor.

Adrenocorticotropic Hormone↗
PubMed1995

Expression of melanocortin 1 receptor in periaqueductal gray matter.

The localization of the melanocortin 1 (MC1) receptor in brain was investigated by in situ hybridization and immunohistochemistry. In rat periaqueductal gray matter (PAG), a small number of cells which displayed a punctate distribution showed a specific hybridization signal for MC1 receptor mRNA, whereas other regions studied were negative. In human PAG, the immunoreactivity for MC1 receptor was detected in scattered cells. These results indicate that a restricted distribution of MC1 receptor is present in the central nervous system.

Animals↗
PubMed1995

Identification of drugs subtype-selective for alpha 2A-, alpha 2B-, and alpha 2C-adrenoceptors in the pig cerebellum and kidney cortex.

The radioligands [3H]MK912 and [3H]RX821002 were used to label alpha2A-, alpha2B-, and alpha2C-adrenoceptors of the pig cerebellum and kidney cortex. By inclusion of the alpha2A-adrenoceptor-selective drug, BRL44408, and using a 'multi-curve' experimental design all the three porcine alpha2-adrenoceptor subtypes could be characterized pharmacologically. The data indicate that the pig alpha2-adrenoceptor subtypes are pharmacologically more related to human alpha2-adrenoceptor subtypes than to the rodent alpha2-adrenoceptors. We suggest a set of drugs that are useful for the delineation of the pig alpha2-adrenoceptor subtypes.

Animals↗
PubMed1995

Dynamic properties of I2-receptors.

Evidence is presented that drug binding to I2-sites in the guinea pig kidney and liver is subjected to dynamic homeotropic allosteric regulation. Drugs have been found that are capable of inducing both positive and negative cooperativity on both kidney and liver I2-sites when studied in competition with [3H]idazoxan. Moreover the kinetics of [3H]idazoxan association with guinea pig kidney I2-sites was complex and occurred with three different rate constants. However, on dissociation of [3H]idazoxan from its binding sites in the kidney a substantial number of the slower sites had been converted to faster sites, thus supporting the idea of a dynamic interconversion of I2 sites. The guinea pig liver and kidney I2-sites appear to be similar in their pharmacological properties. They also bear some resemblance to the I2-sites of the smooth muscle of the guinea pig ileum, but they appear to be more distant to the guinea pig cerebral cortex I2-sites. Nevertheless, data indicate that considerable overall heterogeneity exists between the guinea pig kidney and liver I2-sites and the guinea pig ileum and cerebral cortex I2-sites.

Animals↗
PubMed1995

A three dimensional model for the interaction of MSH with the melanocortin-1 receptor.

A model for the interaction of the melanocortin-1 (MC1) receptor with MSH peptide has been constructed. The model was built by homology modelling using bacteriorhodopsin as template. A cyclic analogue of MSH could be docked into a binding pocket located between transmembrane (TM) domains 2, 3 and 6 of the receptor. The most significant receptor to ligand interactions occur between D117 in TM3 of receptor with histidine in cyclic MSH-peptide, H260 in TM6 with glutamic in peptide and D121 in TM3 with arginine in peptide. The model finds support from earlier mutagenesis data.

Amino Acid Sequence↗
PubMed1995

Polyclonal antibodies against human melanocortin MC1 receptor: preliminary immunohistochemical localisation of melanocortin MC1 receptor to malignant melanoma cells.

Peptides of 11 and 15 residue lengths were synthesised according to the sequence of the N-terminal region of the human MC1 melanocyte stimulating hormone receptor. The peptides were conjugated to thyroglobulin and used for preparation of antisera in the rabbit. Each of the conjugates raised antisera which showed high titre and specificity for its respective peptide antigen when evaluated in an ELISA test. Both types of antisera immunostained MC1 receptor expressing COS-7 cells. By contrast, the sera did not stain control COS-7 cells not expressing the MC1 receptor. Moreover, preimmune sera or antiserum preadsorbed with its respective peptide did not stain the MC1 receptor expressing cells. The antisera were used to immunostain sections of normal human skin, as well as samples of cutaneous malignant melanoma tumours obtained from a patient. The cells of the melanoma tumours were very strongly immunostained with the MC1 receptor antisera. By contrast, melanocytes which were present in the normal skin could not be visualised with our antisera.

Amino Acid Sequence↗
PubMed1995

Characterisation of melanocortin receptor subtypes by radioligand binding analysis.

The DNAs encoding three melanocortin receptor subtypes (melanocortin MC1 receptor, melanocortin MC3 receptor and melanocortin MC5 receptor) were expressed individually in COS (CV-1 Origin, SV40) cells to characterise their ligand binding properties. The results indicated that [125I][Nle4, D-Phe7]alpha-MSH (melanocyte stimulating hormone) bound to a single saturable site with Kd values of 85.1 +/- 8.0 pmol/l (mean +/- S.E.M), 396 +/- 65 pmol/l and 5.05 +/- 1.00 nmol/l for melanocortin MC1 receptor, melanocortin MC3 receptor and melanocortin MC3 receptor, respectively. The melanocortin MC1 receptor and the melanocortin MC5 receptor showed a similar potency order to the melanocortic peptides examined which was markedly different from the potency order of the melanocortin MC3 receptor. The melanocortin MC1 receptor and melanocortin MC5 receptor had a relatively higher affinity for alpha-MSH than gamma-MSH and beta-MSH, whereas the melanocortin MC3 receptor had higher affinity for desacetyl-alpha-MSH, gamma-MSH and beta-MSH compared to alpha-MSH. The inclusion of the endopeptidase inhibitor phosphoramidon to prevent the breakdown of ACTH-(1-39) (adrenocorticotrophic hormone) to alpha-MSH, decreased ACTH-(1-39) binding affinity showing that ACTH-(1-39) had a much lower affinity for melanocortin MC1 receptor than reported earlier.

Adrenocorticotropic Hormone↗
PubMed1995

Comparison of the binding activities of some drugs on alpha 2A, alpha 2B and alpha 2C-adrenoceptors and non-adrenergic imidazoline sites in the guinea pig.

Simultaneous computer modelling of control and guanfacine-masked [3H]-MK 912 saturation curves as well as guanfacine competition curves revealed that both alpha 2A- and alpha 2C-adrenoceptor subtypes were present in the guinea pig cerebral cortex. The Kd value of [3H]-MK 912 determined for the alpha 2A-subtype was 403 pM and for the alpha 2C-subtype 79.8 pM; the receptor sites showing capacities 172 and 19.5 fmol/mg protein, respectively. The Kds of guanfacine were 20 and 880 nM for the alpha 2A- and alpha 2C-adrenoceptor, respectively. In the guinea pig kidney [3H]-MK 912 bound to a single saturable site with Kd 8.34 nM and capacity 285 fmol/mg protein, the site showing pharmacological properties like an alpha 2B-adrenoceptor. Binding constants of 22 compounds for the three guinea pig alpha 2-adrenoceptor subtypes were determined by computer modelling competition curves using for the cerebral cortex a "3-curve assay", for the kidney an "1-curve assay", and using [3H]-MK 912 as labelled ligand. Of the tested drugs guanfacine and BRL 44408 were found to be clearly alpha 2A-selective, Spiroxatrine, yohimbine, rauwolscine and WB 4101, as well as [3H]-MK 912 itself, were found to be alpha 2C-selective. The most selective compounds for alpha 2B-adrenoceptors, when compared to alpha 2A-adrenoceptors, were ARC 239 and prazosin. In the guinea pig kidney [3H]-p-aminoclonidine bound to alpha as well as to non-adrenergic imidazoline sites. The alpha 2-adrenoceptors could be completely blocked using 10 microM (-)-adrenaline without the non-adrenergic sites being affected. During these conditions the analysis of combined saturation and competition studies using labelled and unlabelled p-aminoclonidine with computer modelling revealed that the ligand labelled two different sites with Kds of 310 and 47,000 nM, respectively. Competition curves of 16 compounds for the non-adrenergic [3H]-p-aminoclonidine sites were shallow and resolved into two-site fits. For the high affinity [3H]-p-aminoclonidine site the highest affinities were shown by 1-medetomidine, UK-14,304, guanabenz and detomidine; the Kds of these drugs ranging 26-72 nM. All drugs tested showed low but varying affinities for the low affinity [3H]-p-aminoclonidine site. These data indicated that the [3H]-p-aminoclonidine binding sites of the guinea pig kidney are grossly different from the [3H]-idazoxan binding I2-receptors previously demonstrated also to be present in the guinea pig kidney.

Adrenergic alpha-Antagonists↗
PubMed1993

Molecular cloning of a novel human melanocortin receptor.

A human genomic clone designated MC-2 is isolated. The cloned DNA codes for a protein of 325 amino acids which possesses seven hydrophobic segments, a characteristic of G-protein coupled receptors. The MC-2 receptor is expressed in brain tissue but not in the melanoma cells. When the MC-2 DNA is expressed in COS-7 cells, it binds [125I]-labelled [Nle4, D-Phe7]- alpha melanocyte stimulating hormone (NDP-MSH) which then could be displaced by melanotropic peptides alpha-MSH, beta-MSH, gamma-MSH and adrenocorticotropic hormone, but not by non-melanotropic peptide beta-endorphin. The highest affinity of 5.18 nM was for the NDP-MSH peptide. The novel MC-2 receptor and the MC-1 receptor, described earlier by us (8) showed identical order of affinity for the melanocortin peptides, but the affinities and the fold differences in the affinities to the melanocortin peptides were different when compared to the earlier described MC-1 receptor. The results suggest that the MC-2 DNA codes for a novel melanocortin receptor.

Amino Acid Sequence↗
PubMed1993

Functional expression of rat alpha 2B-adrenoceptor in Escherichia coli.

Rat alpha 2B-adrenoceptor was expressed in Escherichia coli using 'ATG vector' containing cDNA encoding the 'non-glycosylated rat alpha 2-adrenoceptor' (RNG alpha 2). The highest receptor binding activity (using the alpha 2-adrenoceptor ligand [3H]MK 912) was found when transfected bacteria cultures were grown at 30 degrees C for about 4 h after induction with isopropyl beta-D-thiogalactopyranoside (IPTG). Saturation experiments showed that the radioligand bound to a single saturable site with a Kd of 1.42 +/- 0.09 nM and capacity of 281 +/- 6 fmol/mg protein. Binding constants of 14 compounds for the rat alpha 2B-adrenoceptor expressed in E. coli were determined and compared to the values previously obtained for the rat alpha 2B-adrenoceptor when expressed in COS cells as well as for native neonatal rat lung alpha 2B-adrenoceptors. The results indicate that when the rat alpha 2B-adrenoceptor is expressed in E. coli it retains identical ligand binding properties to those found when the receptor is present in the eukaryotic system. Expressing alpha 2B-adrenoceptors in E. coli would, therefore, seem to constitute a valid alternative in, e.g., drug screening and structure analysis of the alpha 2B-adrenoceptors.

Adrenergic alpha-Antagonists↗
PubMed1993

The effect of intrathecal guanfacine and clonidine on the flexor reflex in rats with intact and sectioned sciatic nerves.

We examined the effect of intrathecal (i.t.) administration of the non-selective alpha 2-adrenoceptor agonist clonidine and the alpha 2A-adrenoceptor selective agonist guanfacine on flexor reflex excitability in decerebrate, spinalized, unanesthetized rats before and after unilateral section of the sciatic nerve. Both guanfacine and clonidine dose dependently depressed the flexor reflex in rats with intact nerves. There was a dramatic increase in the sensitivity of the flexor reflex to the depressive effect of i.t. clonidine 4 to 18 days after sciatic nerve section. In contrast, the sensitivity to i.t. guanfacine increased only slightly. The present results suggest that the markedly increased sensitivity to the analgesic effect of the non-selective alpha 2-adrenoceptor agonist does not involve up-regulation of alpha 2A-adrenoceptors. Since radioligand binding data suggested that about 4% of spinal cord alpha 2-adrenoceptors are of the alpha 2C type, whereas alpha 2B-adrenoceptors are not detectable, alpha 2C-adrenoceptors seem to be a good candidate to mediate the up-regulated response to clonidine in axotomized rats. Thus, alpha 2C-adrenoceptor agonists may be useful in the treatment of neuropathic pain.

Analgesia↗
PubMed1993

Evidence for the existence of two forms of alpha 2A-adrenoceptors in the rat.

The alpha 2A-adrenoceptors in rat spleen, kidney, spinal cord and cerebral cortex were studied using [3H]-RX821002 radioligand binding. In the spleen, spinal cord and cerebral cortex, the ligand bound to saturable sites with a Kd of about 1 nmol/l and capacities of 134, 240 and 290 fmol/mg protein, respectively. Computer modelling competition curves for 39 drugs, including those for alpha 2A-, alpha 2B- or alpha 2C-adrenoceptor selective drugs, indicated that the sites labelled by [3H]-RX821002 in the spleen consisted of a single population of alpha 2A-adrenoceptors. However, the competition curves for guanoxabenz were definitely biphasic and resolved into two site fits, indicating that guanoxabenz was binding to both high affinity (Kd = 35 nmol/l) and low affinity (Kd = 8900 nmol/l) alpha 2A-adrenoceptor sites in the proportions 57% and 43%, respectively. The KdS for a number of alpha 2-adrenoceptor subtype selective drugs, measured in competition with [3H]-RX821002 in cerebral cortex and spinal cord, were highly correlated with those obtained in the spleen indicating their alpha 2A-adrenoceptor nature. However, by contrast to the results with the spleen, the guanoxabenz competition curves for the spinal cord and cerebral cortex were monophasic and resolved only into one site fits, the Kd of guanoxabenz being about 4000 nmol/l for both tissues. Drug KdS for kidney alpha 2A-adrenoceptors were also determined using [3H]-RX821002. For nearly all drugs tested, the KdS were highly correlated with those found for the alpha 2A-adrenoceptors in the other rat tissues.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Antagonists↗