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Biomedical subjects

K M Gautvik

Publications and source records attributed to K M Gautvik.

At least 127 records · Page 7Linked to original sources

Sodium and calcium action potentials in cells derived from a rat medullary thyroid carcinoma.

Primary monolayer cultures were made from a rat medullary thyroid carcinoma. All the cultures produced calcitonin throughout the 4 week experimental period. Two different parenchymal cell types could be distinguished, based on a marked size difference. Only the group of smaller cells was electrically excitable. The membrane potential and resistance of these cells were 55.3 +/- 15.4 mV and 623 +/- 275 M omega (mean +2- S.D., n = 23), and current injections produced regenerative responses in all the cells. All-or-none action potentials were recorded in both normal and Na+-free solution. In Na+-free solution the maximum rate of rise of the action potential was reduced by about 70%, and the action potential was reversibly blocked by D-600. It is concluded that the action potentials have both a Na+ and a Ca2+ component, and circumstantial evidence suggests that our recordings were obtained from the calcitonin-producing cells.

Action Potentials↗

Effect of dietary calcium on secretion of calcitonin and gastrin in rats with medullary thyroid carcinoma.

UNLABELLED: Rats transplanted with medullary thyroid carcinoma (MTC) were divided into 3 groups which received for 34 weeks diets with low (0.09%), medium (0.40%) or high (1.80%) calcium content with a calcium/phosphate ratio 2:1. After 6 weeks the different calcium regimens produced corresponding variations in serum calcium and inverse changes in serum magnesium. No alterations in serum proteins were observed. Serum immunoreactive calcitonin (iCT) was unaltered until week 28 of the experiment, but thereafter increased rapidly by about 4-fold. No difference in serum iCT was observed between the 3 groups at any time. Immunoreactive serum gastrin was reduced in rats on medium and low calcium diet compared to those on a high calcium diet. After 28 weeks, when serum iCT was rising, gastrin values fell in rats on high calcium diet to levels approaching those in the other groups. With increased serum iCT (weeks 28-34) serum gastrin concentrations were low and equal in the 3 groups despite different serum calcium concentrations. Electronmicroscopic examination of tumors from the various diet groups did not reveal significant ultrastructural differences. IN CONCLUSION: low, medium and high calcium intake gave corresponding changes in serum calcium. Diet-induced hypercalcaemia was not normalized by elevated serum iCT. In contrast, serum immunoreactive gastrin maintained by high calcium diet was suppressed by hypercalcitonaemia.

Animals↗

Effect of different calcium intakes on bone mineral composition in rats with a calcitonin secreting tumour.

The effects of hypercalcitoninaemia and different calcium intakes on bone composition were studied in rats with and without a transplanted calcitonin secreting tumour (MCT). The serum concentrations of immunoreactive calcitonin (iCT) increased progressively after transplantation. After 32 weeks the serum iCT concentrations in MCT rats were about 5-fold higher and serum ionized calcium was significantly reduced compared to control animals. In rats with hypercalcitoninaemia eating a standard calcium (0.9%) diet for 32 weeks, the dry weight and calcium and sodium content of the long bones were reduced compared to control rats without tumours or rats with a prolactin secreting tumour. The feeding of low (0.09%), medium (0.4%) and high (1.8%) calcium diets to MCT-rats resulted in corresponding levels of total and ionized serum calcium, but there was no difference in iCT concentrations. The MCT-rats taking the low calcium diet had decreased bone mass and calcium content compared to the other dietary groups. In conclusion, rats on normal calcium intake and carrying a calcitonin secreting tumour, but not a prolactin secreting tumour, had reduced femoral dry weight and bone mineral calcium content. Different calcium intake regimens did not affect hypercalcitoninaemia due to a tumour, but parallel changes in femoral dry weight and calcium content were found.

Animals↗

Increase in plasma calcitonin following femoral fracture in rats.

The secretion of calcitonin (CT) has been studied in male rats following a standardized, left femoral fracture after administration of subcutaneous fluanisone and fentanyl citrate anaesthesia. Twenty-five minutes after the fracture, the plasma concentrations of CT were increased by about 20 per cent in young and by about 60 per cent in adult rats compared to prefracture levels of the hormone. Three weeks later, plasma CT had decreased and was not significantly different from prefracture levels. Anaesthesia combined with femoral fracture did not influence plasma calcium significantly, whereas the plasma concentrations of calcium increased in young control rats during 35 minutes of anaesthesia alone. Plasma CT, however, remained unchanged in these control rats in the same period. In rats with a transplanted, CT-secreting, medullary thyroid carcinoma, femoral fracture did not alter the already high plasma concentrations of CT. It is suggested that increase in CT secretion is part of a general response to trauma.

Age Factors↗

Isolation of rat submandibula kallikreins by using immunoadsorption chromatography.

A one-step immunoadsorption method for the isolation of glandular kallikreins is described using the immunoglobulin fraction from rabbit anti-(rat glandular kallikrein) serum coupled to CNBr-activated Sepharose 4B. The adsorptions of 125I-labelled kallikrein or unlabelled kallifrein from 100 000 g submandibular gland supernatants were more than 97% complete. The elution of kallikrein from the immunoadsorbent using guanidine hydrochloride gave about 20% yield, which could be increased up to 70% by including 0.5% bovine serum albumin in the elution buffer. The electrophoretic mobility of eluted submandibular 125I-labelled kallikrein or submandibular glandular kallikrein was not altered after affinity chromatography, as judged by conventional polyacrylamide disc-gel electrophoresis or by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. In addition, the specific esterase and the kininogenase activities of isolated submandibular kallikreins were more than 90% of those of the reference enzyme. This procedure, which results in the isolation of immunologically and biologically active submandibular kallikrein, may also be used for purificaton of other glandular kallikreins that show immunological homology.

Animals↗

Effects of thyroliberin and 4-aminopyridine on action potentials and prolactin release and synthesis in rat pituitary cells in culture.

Effects of thyroliberin (TRH) and 4-aminopyridine (4AP) were studied on prolactin (PRL) secreting rat pituitary tumour cells in culture (GH3 cells). Intracellular recordings obtained from the same cell before and during TRH stimulation showed this peptide to increase the spontaneous firing frequency and prolong the Ca2+ dependent action potentials. These effects were mimicked by 4 AP, which acts by interfering selectively with voltage dependent ionic channels without affecting resting membrane properties. Optimal doses of TRH and 4AP approximately doubled the release of PRL. In contrast, TRH increased PRL synthesis 1.9-fold while 4AP had no effect. Increased PRL synthesis is thus not a direct consequence of the hormone release. We conclude that TRH and 4AP both stimulates PRL release via the facilitating effects on the action potentials. TRH has additional intracellular effects which lead to increased synthesis of the hormone. The effects of TRH responsible for stimulation of PRL synthesis are not causally related to the impulse activity of the surface membrane of the cell.

Action Potentials↗

Intraglandular transport of 125I-glandular kallikrein in the rat submandibular salivary gland.

The transport of radiolabelled rat submandibular gland kallikrein was studied after local administration to the resting and activated rat submandibular gland. The iodinated kallikrein was electrophoretically, immunologically, and biologically indistinguishable from the intact enzyme. After intraductal and intraglandular application the radioactivity in venous effluent was quantitated and characterized. As judged by gel-filtration 125I-kallikrein in venous effluent eluted at a position similar to that seen when the iodinated enzyme was mixed with plasma, but earlier than the elution of 125I-kallikrein in buffer. In plasma, therefore, glandular kallikrein is probably bound to macromolecules. The radioactive fractions in venous effluent did not contain free iodine. Maximum concentration of 125I-kallikrein in venous effluent of resting glands was repeatedly reached about 20 min after intraductal administration. Moreover, the ductal epithelium represented the main permeation barrier since after intraglandular application the maximum venous 125I-kallikrein concentration was reached almost immediately. In activated gland (parasympathetic and sympathetic nerve stimulation), the venous 125I-kallikrein concentration was inversely related to glandular blood flow. We conclude that kallikrein present in the duct lumen or in the interstitium is able to reach the circulation, thereby making possible the local generation of plasma-kinins.

Animals↗

Plasma concentration of secretin and gastric secretion following intraduodenal calcium infusion in man.

The aim of the present study was to test whether the previously demonstrated release of immunoreactive plasma secretin following intraduodenal bile infusion could be mediated by the calcium content of the bile. The gastric acid secretion and the serum concentration of gastrin, ionized and total calcium, calcitonin, and parathyroid hormone were also measured. A total amount of 93 mg calcium was infused intraduodenally in eight healthy volunteers during constant gastric aspiration. No effect was seen on the concentration of immunoreactive plasma secretin or serum gastrin levels in peripheral blood. The total serum calcium concentration decreased slightly during the infusion and increased immediately thereafter. Ionized serum calcium increased throughout the experiment, the increase being significant only after the calcium infusion was stopped. No effects were observed on the serum concentrations of calcitonin and parathyroid hormone.

Adult↗

The use of region specific radioimmunoassays for characterization of circulating calcitonin in patients with medullary carcinoma of the thyroid gland.

UNLABELLED: Two antisera with known region specificities have been used to characterize calcitonin immunoreactivity (iCT) in serum of patients with medullary thyroid carcinoma (MCT). Antiserum I which was raised against the synthetic hormone (1-32 amino acid residues), contained heterogeneous populations of immunoglobulins directed predominantly against carboxy-terminal sequences of the hormone, but the antiserum reacted also with the amino-terminal fragment (1-10 amino acid residues). Antiserum II, which was raised against the carboxy-terminal hormone fragment (11-32 amino acid residues) reached equally well with the intact hormone and the C-terminal fragment, but showed negligible binding of the amino terminal fragment. Antiserum I measured therefore both amino-terminal and carboxy-terminal sequences of calcitonin while antiserum II measured only carboxy-terminal amino acid sequences. In 40 patients with MCT, antiserum I measured usually the highest concentration of serum iCT suggesting the presence of non-uniform hormone immunoreactivity. The different molecular forms of circulating iCT in 7 MCT patients were explored by using antiserum I after gel filtration on Sephadex G-100. The patients who were selected on basis of iCT measurement in serum using antiserum I and II, could be divided into 3 groups which showed characteristic iCT profiles. Group 1, in which antiserum II measured a higher concentration of serum iCT, contained predominantly (60-70%) small fragments of calcitonin immunoreactivity. On the other hand, in the sera of group 3 in which antisera I measured an equal or the highest concentrations, the dominant form of the hormone consisted of molecular sequences equal to or larger than the intact hormone (90%). In group 2, the two antisera measured an equal amount of serum iCT and molecular forms consisting mostly of larger hormone fragments dominated (50%). All the patients were normocalcaemic in spite of frequently grossly elevated serum iCT, and 33 out of 36 patients had normal serum immunoreactive parathyroid hormone. IN CONCLUSION: 1. Serum iCT is heterogeneous and represents peptides of quite different molecular size with no or low biological activity. 2. Most of the serum calcitonin immunoreactivity consists of peptides with carboxy-terminal amino acid sequences. 3. Most, if not all, of the amino-terminal calcitonin immunoreactivity is due to monomeric and polymeric hormonal forms.

Adult↗

Development of sequence specific radioimmunoassay of human parathyroid hormone and its use in the diagnosis of hyperparathyroidism.

Two antisera which were raised against bovine parathyroid hormone (bPTH), and which cross-reacted with the human hormone, have been characterized. The antisera which originated from rooster and guinea-pig, were found to contain several populations of antibodies directed against both N-terminal and C-terminal sequences of the hormone. However, at proper dilutions the rooster antiserum did not bind the N-terminal fragment nor could this fragment displace the [125I] bPTH (1--84 amino acid residue) from binding to the antiserum. Furthermore, preincubation experiments with excess N-terminal fragment showed only a negligible reduction in maximal binding of the iodinated intact hormone using the rooster antiserum. In contrast, the guinea-pig antiserum reacted equally well with the N-terminal fragment and the intact hormone, and preincubation with this fragment reduced the binding of the [125I]bPTH (1--84 amino acid residues) by 75%. Gel filtration of hyperparathyroid serum on Bio-Gel P-60 showed immunoreactive material which was measured with both antisera, eluting at a position similar to the intact hormone. However, in the C-terminal specific, but not in the N-terminal specific radioimmunoassay the major component eluted together with or somewhat earlier than the N-terminal bPTH fragment (1--34 amino acid residue), and this peak represented more than 90% of total immunoreactive PTH (iPTH) in serum. This major iPTH component must therefore represent fragment(s) with intact carboxy-terminal sequences. The N-terminal specific radioimmunoassay was unable to measure iPTH in about 80--90% of healthy individuals while the C-terminal specific assay detected iPTH in about 88% of these sera (equal to or above 0.1 micrograms/l). Similarly, the N-terminal specific antiserum measured consistently lower serum iPTH concentrations in patients with primary hyperparathyroidism. In thirty-four out of forty-one patients with surgically verified primary hyperparathyroidism, serum iPTH concentrations equal to or above 0.60 micrograms/l were demonstrated using the C-terminal, specific radioimmunoassay.

Acute Disease↗

Receptors for 17beta-estradiol in prolactin-secreting rat pituitary cells.

Estrogens stimulate prolactin (PRL) synthesis by GH3 cells, a clonal strain of rat pituitary cells grown in culture. At 4 degrees C the binding of [3H]17 beta-estradiol to monolayer cultures of GH3 cells was specific and of limited capacity, with half-maximal and maximal binding after 1--2 h and 12 h, respectively. Scatchard analysis showed one single class of binding sites with Kd = 3.1 X 10(-10) M and n = 309 X 10(-15) mol 17 beta-estradiol/mg cell protein, calculated to give approx. 25,000 binding sites per cell. At 4 degrees C less than 10% of the specifically bound [3H]17 beta-estradiol was found in the nuclear fraction. When the incubation temperature was raised to 37 degrees C, the amount of radioactivity in the nucleus increased to 25% within 30 min with a corresponding reduction in the cytoplasm. The cytosol fractions from monolayer cultures as well as from tumors of GH3 cells contained specific 17 beta-estradiol binding proteins, having a sedimentation constant close to 8S in a salt-free buffer and 4S in the presence of 0.5 M KCl. scatchard analysis showed one single class of binding sites with Kd = 3.6 X 10(-10) M and n = 258 X 10(-15) mol 17 beta-estradiol/mg cytosol protein (GH3 tumor tissue). Thus, GH3 cells grown in culture and in the intact animal have similar binding characteristics as judged from the data for binding affinity, capacity and specificity. After the in vivo administration of [3H]17 beta-estradiol to GH3 tumor-bearing rats, radioactivity could be extracted (0.5 M KCl) from purified nuclei bound to 4.5S macromolecules. We suggest that the action of 17 beta-estradiol on GH3 cells involves an initial binding of the steroid to specific receptors in the cytoplasm, followed by transport of a fraction of the hormone-receptor complexes to the nucleus involving a temperature-sensitive step.

Animals↗