Cary W. Cooper

Provocative Agents and the Diagnosis of Medullary Carcinoma of the Thyroid Gland

Twenty-six patients with known or suspected medullary thyroid carcinoma (MTC) and 21 normal control subjects were tested intravenously on four separate days with calcium gluconate (CG), 2 mg Ca++/kg/l min.; pentagastrin (P), 0.5 ug/kg/ 5 sec; calcium chloride (CC), 3 mg Ca++/kg/10 min.; and a combination of calcium gluconate and pentagastrin (CG + P). Calcitonin (CT) levels were determined by radioimmunoassay on plasma collected before and immediately following each test infusion. In none of the 21 control subjects was there a clear increase in CT above 200 pg/ml following any of the four provocative tests. Conversely, in all 26 patients with known or suspected MTC, plasma CT levels were markedly increased (>300 pg/ml) following the combined infusion of CG + P. The peak CT response was greater with CG + P than with a) CG alone (22 of 24 patients, p < 0.002), b) P alone (25 of 26 patients, p < 0.002), or c) CC alone (17 of 17 patients, p < 0.002). Of 12 MTC patients with undetectable basal calcitonin levels, all had peak responses greater than 300 pg/ml following CG + P, whereas such responses occurred less often following CG alone (8 of 12) or P alone (8 of 12). The results demonstrate that the combined administration of pentagastrin and calcium gluconate constitutes a more effective and reliable stimulus for CT secretion from MTC cells than the use of either agent alone, and appears the most useful single screening test for the detection of occult MTC.

Thyrocalcitonin: Stimulation of Secretion by Pentagastrin

Administration of a small dose of pentagastrin, a synthetic pentapeptide containing the biologically active portion of the native hormone gastrin, results in a marked, rapid, transitory increase in thyrocalcitonin secretion in the pig. Gastrin or a related gastrointestinal peptide may be important in the physiological secretion of thyrocalcitonin, such as that which occurs when calcium salts are introduced into the gastrointestinal tract.

Medullary thyroid carcinoma: Relationship of method of diagnosis to pathologic staging

Medullary thyroid carcinoma (MTC) develops in virtually all patients affected with multiple endocrine neoplasia type II (MEN II), a disease inherited as an autosomal dominant trait. The thyroid tumor cells secrete calcitonin (CT) and the detection of elevated plasma levels (>300 pg/ml) of this hormone in MEN II kindred members strongly suggests the presence of MTC even though it may not be evident clinically. Intravenously administered calcium ion (Ca++) and pentagastrin (Pg) are potent CT secretagogues which are of particular value in establishing the early diagnosis of MTC. In evaluating seven kindreds with MEN II, we detected 90 patients with MTC. Depending on the method of diagnosis, they could be divided into three categories: Group 1; patients with no clinical evidence of MTC whose undetectable basal plasma calcitonin levels became elevated following intravenous Ca++ or Pg, Group II; patients with no clinical evidence of MTC who had elevated basal plasma CT levels, and Group III; patients with clinically evident MTC. At the time of diagnosis of MTC, the patients in Group I were younger (20.5 ± 1.9 years) than the patients in Group II (32.5 ± 4.7 years, p < 0.005) and Group III (34.3 ± 2.0, p < 0.00005). The incidence of residual MTC, as indicated by an elevated plasma CT level following provocative testing post operatively, was less frequent in patients diagnosed biochemically ([6/34]; Group I, 4/26 and Group II, 2/8) than in those diagnosed clinically (Group III, 15/26, p<0.002). Furthermore, regional nodes were involved less often in patients diagnosed biochemically ([5/28]; Group I, 2/22 and Group II, 3/6) than in those diagnosed clinically (Group III, 15/24, p < 0.02). Distant metastases were only evident in Group III patients. Patients with MEN II who had the diagnosis of MTC established biochemically rather than clinically, had a more favorable pathological stage of disease at the tune of thyroidectomy. This was especially true if the biochemical diagnosis had been by provocative testing.

Ability of calcitonins to alter food and water consumption in the rat.

Synthetic salmon calcitonin (sCT) given SC (30 MRC U/kg) or ICV (1.4 U, 300 ng) inhibited 24-hr food consumption in the rat by 50-100%. Furthermore, ICV doses of sCT ranging from 75 ng (0.35 U) to 300 ng (1.4 U) lowered both food and water intake in a dose-dependent manner. Use of various forms of CT give an apparent order of potency of salmon greater than porcine greater than or equal to human with sCT being at least 40 fold more potent than the mammalian forms. Measurement of food and water intake over a brief (30-90 min) period showed that ICV sCT was effective in reducing food and water consumption regardless of whether it had been given 1, 12, or 23 hr previously. Daily administration of sCT for 5 days caused marked suppression of food and water intake for 2 days; thereafter, consumption returned toward normal, becoming equivalent to vehicle injected controls by day 3 or 4 despite continued daily injections of sCT. The results show that CT can act centrally to modify certain types of behavior and are of special interest since CT-like peptides have been described in the pituitary and hypothalamus and since CT receptors have been reported in hypothalamic and other brain regions.

Relationship of Blood Concentrations of Calcium, Phosphate, Gastrin and Calcitonin to the Onset of Feeding in the Rat

Summary Daily fluctuations in plasma calcium concentrations in rats trained to a closely regulated feeding pattern have been compared to corresponding plasma gastrin and calcitonin concentrations. The time period studied was that extending from 4 hr prior to the start of the feeding period through the first 2 hr of feeding. Both plasma calcium and phosphate levels fell prior to the start of the feeding period and remained low at least for the first 2 hr of feeding. This pattern was also observed in rats in which food was withheld for 2 hr past the regular feeding time. Plasma 45Ca and 32P concentrations (radionuclide injected at least one week prior to sampling) did not follow the pattern of their stable counterparts. Instead, these values rose or remained constant until after feeding had commenced, after which they fell precipitously. Both plasma calcitonin and gastrin levels rose rapidly after the start of the feeding period. The primary point of emphasis is that calcitonin secretion was produced in these rats by an intestinal related stimulus and not by a rise in plasma calcium concentration. The technical assistance of Dorothy B. Raneri, Blanche P. Holloway and Johnny F. Obie is acknowledged and appreciated. Thanks are expressed to Merlene Moore for clerical assistance.

Calcitonin and calcitonin gene-related peptide enhance calcium-dependent potentials.

Recent data suggests that calcitonin (CT) and/or calcitonin gene-related peptide (CGRP) may be potential transmitters or modulators in the nervous system. The present study analyzed the effect of CT and CGRP on the neuronal membranes of cat parasympathetic ganglia of the urinary bladder. The related peptides prolonged the duration of the afterhyperpolarization of the action potential but had no effect on resting potential or input resistance. CT and CGRP enhanced the duration of a calcium spike recorded in the presence of agents blocking Na and K channels while under similar conditions forskolin, an activator of adenylate cyclase, did not affect the calcium spike. These data suggest that the neural mechanism of action of CT and CGRP is to prolong a calcium conductance and that these effects are not mediated through cyclic AMP.

Colocalization of calcitonin gene-related peptide and somatostatin in pancreatic islet cells and inhibition of insulin secretion by calcitonin gene-related peptide in the rat.

Calcitonin gene-related peptide (CGRP)- and somatostatin (SR1F)-containing cells were identified by immunocytochemical techniques in pancreatic islet cells of the rat. CGRP-containing cells were found primarily in the peripheral portion of the pancreatic islets. In addition, CGRP-containing cells also contained somatostatin, which identifies the islet CGRP-containing cells as D cells. In the present study, we also tested the effect of CGRP on gastrin-releasing peptide (GRP; M)-stimulated release of insulin from isolated rat islets in vitro. At concentrations of 10−8 – 10−8 M, CGRP inhibited GRP- and CCK-8-stimulated release of insulin significantly when compared with GRP or CCK-8 alone. At the lowest concentration of CGRP (10−8 M). the inhibitory effect of CGRP on CCK-8-stimulated release of insulin was statistically significant (p <0.05) and exceptionally potent (65–90% inhibition). We have also found that CGRP does not stimulate the release of SRIF from isolated islet cells. These findings suggest that CGRP may play a regulatory role in the release of insulin.

Endogenous parathyroid hormone-related peptide enhances proliferation and inhibits differentiation in the osteoblast-like cell line ROS 17/2.8.

To investigate potential effects of endogenous parathyroid hormone-related peptide (PTHrP) on osteoblast function, ROS 17/2.8 cells were transfected with full-length PTHrP cDNA in a sense or antisense orientation to alter PTHrP production. Compared with vector-transfected control cells, PTHrP-overproducing (sense-transfected) cells showed increased DNA synthesis ([(3)H]-thymidine incorporation) and increased growth (cell number). The extent of apoptosis was compared for the different clones using the terminal deoxynucleotide-mediated dUTP nick-end-labeling assay (TUNEL) and Hoechst staining. No differences in percentages of apoptotic cells were found under basal culture conditions or after 3 days of serum deprivation, which, itself, markedly increased numbers of apoptotic cells. The effect of PTHrP on osteoblast differentiation was assessed by examining two protein markers of differentiation, alkaline phosphatase, and bone morphogenetic protein (BMP)-2. Alkaline phosphatase activity was decreased in sense-transfected cells and increased in antisense-transfected cells, compared with cells transfected with empty vector. PTHrP-overproducing cells also showed decreased numbers of BMP-2-positive cells, whereas antisense-transfected cells showed no difference compared with vector control. The results indicate that: (a) endogenously produced PTHrP can increase growth of these osteoblastic cells by stimulating proliferation while not affecting apoptosis; and (b) the increased cell proliferation produced by PTHrP was accompanied by a reduction in activity or amount of two proteins normally expressed by differentiated osteoblasts.

Regulation of Calcium Transport in Bone by Parathyroid Hormone

Publisher Summary This chapter discusses regulation of calcium transport in bone by parathyroid hormone. It attempts to review current concepts concerning the mode of action of parathyroid hormone, and points out apparent similarities between in vivo effects of this hormone on a variety of metabolic functions and those produced by elevated circulating calcium concentrations. It also presents a model to account for parathyroid control of calcium fluxes in bone. In this chapter, it is proposed that by enhancing entry of calcium into the interior of the cell, parathyroid hormone produces two divergent effects on the overall metabolism and physiology of bone and other target tissues. The first effect is that by allowing increased intracellular ionic calcium levels, the hormone increases the amount of calcium transported across specific layers of target cells. This net movement of calcium is unidirectional or “polarized” and occurs against a concentration gradient via a “pump” mechanism. The second is that if the intracellular concentration of ionic calcium is appreciably increased, many enzymatically controlled processes within the cell may, in turn, be affected. This secondary function of parathyroid hormone can be termed “stimulation of calcium-influenced metabolic activities” and may occur in any cell that responds to the hormone by allowing increased entry of calcium.

Calcitonin depresses amphetamine-induced locomotor activity

Synthetic salmon calcitonin (sCT), given subcutaneously (6.4 micrograms/kg) or intracerebroventricularly (ICV, 30-600 ng), depressed amphetamine-induced locomotor activity in rats by more than 50%. ICV injection of sCT, either three hours or immediately before intraperitoneal amphetamine (1-3 mg/kg), significantly reduced the amphetamine-induced activity. In the absence of amphetamine, sCT had no effect on locomotor activity during the first 100 minutes after treatment. These results show sCT can act centrally to modify drug-induced behavior and may be related to reports of calcitonin receptors and calcitonin-like peptides in the brain.