Rat Parathyroid Research Articles

Blood vascular bed and pericapillary space in rat parathyroid glands

The blood vascular bed and pericapillary space of the rat parathyroid gland were studied by scanning electron microscopy of vascular casts, freeze-cracked tissue blocks, and NaOH-treated tissue specimens. The findings were supplemented by transmission light and electron microscopy of sectioned tissue samples. The rat parathyroid gland contained a rich network of freely anastomosing capillaries. These capillaries were surrounded by marked pericapillary spaces that were demarcated by basal lamina of both capillaries and parenchymal cells. The pericapillary spaces contained many collagen fibrils and frequently issued some projections running deep into the sheets of parathyroid cells. The latter projections may be useful to supply the parenchymal cells located far from the capillaries. The collagen fibrils may regulate the flow of tissue fluid in the pericapillary space and convey parathyroid hormone, which is released at the apicolateral domain, into the capillaries.

Quantitative and three‐dimensional aspects of the rat parathyroid gland in normo‐, hypo‐, and hypercalcemia

The ultrastructure of the rat parathyroid has been under study for more than 35 years, but controversies still exist, especially regarding structure-function relationships. The present review focuses on recent morphological parathyroid research on rats under normal conditions and in various states of disturbed calcium metabolism. To facilitate discussions on functional aspects, current biochemical data, particularly those dealing with the regulation of parathyroid hormone synthesis and release, are also considered. Our results from quantitative studies and from investigations employing serial sectioning form the basis for the discussions. A central issue is whether the parathyroid secretory cells undergo secretory cycles. Prompted by results obtained from improved fixation procedures and serial sectioning, we question the basis for the theory of secretory cycles. Since the rat parathyroid secretory cell is polar, a single section is not an appropriate sample for estimating functional activity and for comparing the structure and distribution of intracellular components of adjacent cells. The heterogeneity in ultrastructural appearance of intracellular vesicles calls for the use of specific markers in relating the structure of the vesicular compartment to intracellular processing of hormone. The importance of unbiased quantitative techniques is illustrated in discussions on cell number and size for estimating the response of the parathyroid gland to different functional states or disorders demanding changes in secretion of parathyroid hormone, e.g., hyper- and hypocalcemia.

Mammalian parathyroids: Morphological and functional implications

Fixation with aldehydes is achieved either by immersion or perfusion. The parenchyma of parathyroid glands fixed by immersion consists of dark cells containing a lot of membranes of these organelles which are concerned with hormone secretion, light cells which are poor in these organelles, intermediate forms between the two, and multinuclear syncytial cells. They have been attributed to represent different functional stages of secretory activity, the dark cell being in an active form, the light cell in a resting form. Studies of the parathyroids of mice, rats, rabbits, cats, dogs, pigs, cattle, sheep, goats, and horses employing various fixation protocols clearly demonstrate that light cell variants and multinuclear syncytial cells are formed during improper immersion fixation as a result of membrane disintegration. Parathyroids fixed by perfusion or by immersion in an appropriate fixation medium comprise only one cell type which correspond to the dark chief cell. Parathyroid cells are polar cells bearing some of the rough endoplasmic reticulum in the basal pole, the rest of it, the Golgi complex, and secretory granules in the apical pole. The secretory product is released by exocytosis at the apicolateral domain of the plasma membrane into the intercellular space. Secretory activity can be altered experimentally, leading to drastic changes in the amount of cell membrane related to hormone synthesis, intracellular transport, exocytic release, and secretion coupled membrane retrieval. The sensitive reaction of parathyroid cells to both the mode of fixation and to fixation media demands careful evaluation of the fixation protocol. This and the polarity of parathyroid cells have to be borne in mind for estimating secretory activity on the basis of morphological criteria.

Microvascularization of corpuscles of Stannius in teleost fishes.

Blood supply and microvascular patterns of Stannius corpuscles were studied by scanning electron microscopy of vascular corrosion casts in the teleost fishes Blennius pavo, Zosterisessor ophiocephalus, and Gasterosteus aculeatus. Microvascular casts demonstrated that Stannius corpuscles--depending on their location--have an arterial supply derived either directly from the dorsal aorta, from the trunk of the first ventral segmental artery of the tail, or from a renal artery. Supplying arteries form a capsular capillary bed and a parenchymal capillary bed; both are composed of fine, freely anastomosing vessels with a homogeneous isotropic distribution. Central venules arise deep in the corpuscles. In the capsule, they form a single vein which drains into a segmental vein or directly into the caudal vein. Stanniocalcin, the hormone of the Stannius corpuscle, enters the renal circulation and reaches its main target organs, the gills, via posterior cardinal veins--heart--ventral aorta. Occasionally, some capsular venules empty into the trunk kidney peritubular venules. Capillaries are fenestrated and are embraced by pericytes with long, slender processes. The perivascular space contains collagen fibrils. Nerve fibers are found close to endothelial cells and pericytes. Vascular patterns of Stannius corpuscles are compared with those of the rat parathyroid glands and are discussed in respect to physiological implications.

The biological significance of storage granules in rat parathyroid cells.

Both prosecretory and storage granules are concomitantly formed at the trans Golgi network including the innermost Golgi cisterna. Prosecretory granules develop into small secretory granules that release their contents by exocytosis finely regulated by a complex mechanism for maintaining calcium homeostasis. In the rat parathyroid cells, storage granules are large secretory granules storing parathyroid hormone for an emergency supply. The hormone is rapidly discharged by exocytosis when serum calcium concentration is decreased. The granules are constantly produced even under conditions of low serum calcium concentration in the regions of 8 mg/dl. The granule content is constantly hydrolyzed when not discharged, leading to a decreased core and finally to the formation of vacuolar bodies. The fate of the vacuolar bodies is unknown. Hypercalcemic conditions accelerate hydrolysis. The threshold value of calcium concentration required for the release of storage granule contents is between 8.0 and 7.5 mg/dl and that of calcium concentration for accelerating degradation of storage granules is about 11.5 mg/dl. Sympathetic stimulation causes storage granules to be discharged regardless of hypercalcemia or hypocalcemia. Parasympathetic stimulation accelerates hydrolysis. The degradation of storage granules seems to be closely associated with an intracellular regulatory mechanism for parathyroid hormone secretion.

Effect of Parathyroid Hormone and Calcitonin on Cholinergic Markers in Rat Parathyroid Gland

The effect of parathyroid hormone (PTH) and calcitonin on 3H-choline uptake and on 3H-acetylcholine synthesis by rat parathyroid glands were examined. Incubation of tissue for 120 min with PTH or calcitonin resulted in an inverted bell-shaped, dose-dependent inhibition of 3H-choline uptake, with a maximal effect at 10(-9) M concentration. The effect of PTH on parathyroid choline uptake was blunted by preincubation with the PTH antagonist NLe(8-18)-PTH (3-34) amide. PTH brought about a dose-dependent inhibition of 3H-choline conversion to 3H-acetylcholine, with significant effects at 10(-8) M concentration or higher. This inhibitory effect of PTH on 3H-acetylcholine synthesis was blocked by co-incubation with the PTH antagonist NLe(8-18)-PTH (3-34) amide. Only at a 10(-7) M concentration calcitonin was effective to impair the in vitro conversion of radioactive choline into 3H-acetylcholine by parathyroid fragments. The results indicate that, in vitro, PTH, and to a less extent, calcitonin, inhibit cholinergic activity in rat parathyroid glands.

The roles of calcium and 1,25-dihydroxyvitamin D3 in the regulation of vitamin D receptor expression by rat parathyroid glands.

Expression of the vitamin D receptor (VDR) in the parathyroid glands is decreased in secondary hyperparathyroidism associated with chronic renal failure by undefined mechanisms. In the present study, we examined the effects of hyperparathyroidism and dietary calcium and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] on the expression of VDR in rat parathyroid glands. Vitamin D-deficient rats were maintained on diets containing 0.02% Ca (-D, LCD), 0.4% Ca (-D, NCD), or 2.0% Ca (-D, HCD) for 6 weeks. Serum ionized Ca (ICa) in the rats on the three diets ranged from 2.5-5.2 mg/dl. Serum PTH ranged from 22-590 pg/ml and correlated inversely with ICa (r = -0.835; P < 0.001). Rats with the highest ICa had normal PTH values, suggesting that vitamin D deficiency per se does not lead to hyperparathyroidism. VDR messenger RNA (mRNA) levels in the parathyroid glands correlated positively with ICa (r = 0.845; P < 0.001) and negatively with PTH (r = -0.716; P < 0.001). VDR mRNA levels in the rats fed the -D, HCD were 6 times higher than those receiving -D, LCD and the same as those in rats fed a normal (Purina) diet. Thus, prevention of hyperparathyroidism with high dietary calcium prevented the drop in VDR expression. Treatment of the rats on all three diets with 0, 25, or 100 ng 1,25-(OH)2D3, ip, 48 and 12 h before death dose dependently increased ICa and decreased PTH, as expected, and also increased parathyroid gland VDR mRNA. This coordinate regulation of VDR mRNA by calcium and 1,25-(OH)2D3 was also observed in the kidney, but intestinal VDR mRNA was not stimulated by dietary calcium or 1,25-(OH)2D3. Analysis of covariance for parathyroid gland VDR mRNA and ICa for the three doses of 1,25-(OH)2D3 revealed no significant independent effect of 1,25-(OH)2D3 on VDR mRNA, suggesting that the up-regulation of VDR expression by 1,25-(OH)2D3 in the parathyroid glands may be mediated primarily by increasing serum calcium.

The roles of calcium and 1,25-dihydroxyvitamin D3 in the regulation of vitamin D receptor expression by rat parathyroid glands

The roles of calcium and 1,25-dihydroxyvitamin D3 in the regulation of vitamin D receptor expression by rat parathyroid glands

Truncation of the Carboxyl-terminal Region of the Rat Parathyroid Hormone (PTH)/PTH-related Peptide Receptor Enhances PTH Stimulation of Adenylyl Cyclase but Not Phospholipase C

The functional role of the rat parathyroid hormone(PTH)/PTH-related peptide (PTHrP) receptor's carboxyl-terminal region was characterized by comparing the binding and signaling properties of receptors that have 78 and 111 amino acid deletions (R513 and R480, respectively), with those of the 591-amino acid wild-type (WT) receptor. R480 and R513 have 4- and 1.5-fold lower apparent Kd values for rat PTH-(1-34) (rPTH), compared with the WT receptor (WT, 1.81 +/- 0.19 nM; R513, 1.24 +/- 0.12 nM; R480, 0.48 +/- 0.05 nM, mean +/- S.E.). PTH (100 nM)-stimulated cAMP accumulation and polyphosphoinositide hydrolysis both correlated positively with receptor expression. However, whereas PTH-stimulated polyphosphoinositide hydrolysis was indistinguishable among WT and either truncated mutant at comparable levels of expressed receptors, maximal PTH-stimulated cAMP accumulation was 4-6- and 2-3-fold higher in cells expressing R480 and R513, respectively. Furthermore, pretreatment of COS-7 cells with 100 ng/ml of pertussis toxin (PTX) enhanced PTH-stimulated cAMP accumulation in cells expressing the WT receptor, but failed to do so in cells expressing either R480 or R513. Thus, sequences in the PTH/PTHrP receptor's carboxyl-terminal tail lower the affinity of the WT receptor for agonist; directly interact with, or indirectly facilitate the interaction of the receptor with a PTX-sensitive G protein that inhibits adenylyl cyclase; and decrease the efficacy with which the receptor interacts with Gs.

Acute Effect of Parathyroid Hormone on Urine Concentration in the Rat

1. It has been demonstrated that parathyroid hormone can increase adenylate cyclase activity in the rat papilla, produce a small antidiuretic effect and in vitro can interfere with the action of arginine vasopressin on water transport. Clearance studies were performed in the anaesthetized water diuretic thyroparathyroidectomized rat to evaluate further the effect of parathyroid hormone on urine concentration in the presence and absence of arginine vasopressin. 2. A maximal phosphaturic concentration of rat parathyroid hormone (2 micrograms/kg) reduced urine flow from 125 +/- 7 to 81 +/- 9 microliters/min within 10 min (P < 0.01). Addition of a maximal antidiuretic concentration of arginine vasopressin (100 ng/kg) produced a delayed and diminished antidiuretic response when compared with a group of rats not pretreated with parathyroid hormone (47 +/- 5 compared with 27 +/- 5 microliters/min; P < 0.01). However, a supramaximal arginine vasopressin concentration (1000 ng/kg) produced a maximal antidiuretic effect in the presence of parathyroid hormone. 3. To evaluate further the inhibitory effect of parathyroid hormone on arginine vasopressin-induced antidiuresis, parathyroid hormone (2 micrograms/kg) was administered to one group of rats and a minimally effective arginine vasopressin concentration (7.5 ng/kg) to another group, which produced a similar antidiuretic effect. However, the subsequent effect of a maximal antidiuretic arginine vasopressin concentration (100 ng/kg) was again significantly blunted in the group pretreated with parathyroid hormone. 4. Parathyroid hormone produced only a small increase in mean plasma calcium concentration, and glomerular filtration rate was not altered by either hormone.(ABSTRACT TRUNCATED AT 250 WORDS)

Induction of bone resorbing-activity by mouse stromal cell line, MC3T3-G2/PA6.

Unfractionated mouse long-bone cells prestimulated with rat parathyroid hormone (PTH) for 7 days (PTH-prestimulated long-bone cells; PTH-psLB cells) contained about 20% tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells and about 0.2% TRAP-positive multinucleate cells. When PTH-psLB cells (1.0 x 10(5)/slice) were cultured on dentine slices, about 100 resorption lacunae (pits) per slice were formed by 6 h after the start of culture, while the number of TRAP-positive multinucleate cells on the dentine slices did not increase. When the number of inoculated cells was plotted versus the number of pits formed during the 6-h cultivation, a sigmoid curve was generated. When PTH-psLB cells (1.0 x 10(3)/slice) were cocultured with various mouse stromal cell lines, the MC3T3-G2/PA6 line was found to increase the number of pits. Since this activity was manifested within a short period of time (4 h), and as there was no change in the number of TRAP-positive multinucleate cells, we postulate that the activity of MC3T3-G2/PA6 is exerted on mature osteoclastic cells capable of resorbing bone rather than on some differentiation step of the osteoclast cell lineage. With respect to the mechanism of expression of this activity, it is unlikely that MC3T3-G2/PA6 produces a soluble factor, because no activity was seen when MC3T3-G2/PA6 and PTH-psLB cells were cultured separately across a membrane in a Transwell. These results suggest that cell-to-cell contact and communication between PTH-psLB cells and MC3T3-G2/PA6 may be a crucial event for the stimulation of osteoclastic bone resorption by MC3T3-G2/PA6.

Induction of bone resorbing-activity by mouse stromal cell line, MC3T3-G2/PA6

Unfractionated mouse long-bone cells prestimulated with rat parathyroid hormone (PTH) for 7 days (PTH-prestimulated long-bone cells; PTH-psLB cells) contained about 20% tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells and about 0.2% TRAP-positive multinucleate cells. When PTH-psLB cells (1.0 × 10 5/slice) were cultured on dentine slices, about 100 resorption lacunae (pits) per slice were formed by 6 h after the start of culture, while the number of TRAP-positive multinucleate cells on the dentine slices did not increase. When the number of inoculated cells was plotted versus the number of pits formed during the 6-h cultivation, a sigmoid curve was generated. When PTH-psLB cells (1.0 × 10 3/slice) were cocultured with various mouse stromal cell lines, the MC3T3-G2/PA6 line was found to increase the number of pits. Since this activity was manifested within a short period of time (4 h), and as there was no change in the number of TRAP-positive multinucleate cells, we postulate that the activity of MC3T3-G2/PA6 is exerted on mature osteoclastic cells capable of resorbing bone rather than on some differentiation step of the osteoclast cell lineage. With respect to the mechanism of expression of this activity, it is unlikely that MC3T3-G2/PA6 produces a soluble factor, because no activity was seen when MC3T3-G2/ PA6 and PTH-psLB cells were cultured separately across a membrane in a Transwell. These results suggest that cell-to-cell contact and communication between PTH-psLB cells and MC3T3-G2/PA6 may be a crucial event for the stimulation of osteoclastic bone resorption by MC3T3-G2/PA6.

Molecular cloning and expression of the acidic fibroblast growth factor receptors in a rat parathyroid cell line (PT-r). Parathyroid cell-specific calcium-dependent change of ligand accessibility and covalent attachment of heparan sulfate glycosaminoglycan to the receptors.

We have previously identified two fibroblast growth factor (FGF) receptors with higher affinity for acidic FGF rather than basic FGF in a rat parathyroid cell line (PT-r). Carbohydrate analyses of the receptors suggested the presence of three different types of FGF receptors, a 150-kDa glycoprotein receptor, a approximately 150-kDa heparan sulfate-proteoglycan receptor, and a 130-kDa glycoprotein receptor (Sakaguchi, K., Yanagishita, M., Takeuchi, Y., and Aurbach, G.D. (1991) J. Biol. Chem. 266, 7270-7278). Here, we have cloned two isoforms of the FGF receptors from PT-r cells; one with two immunoglobulin (Ig)-like domains (clone a), and the other with an additional Ig-like domain and an acidic box (clone b). They showed highest homology to the mouse and human keratinocyte growth factor receptors among the FGF receptors reported. Clones a and b had one and three possible glycosaminoglycan attachment sites, respectively. Heparitinase treatment of PT-r cells transfected with clone a suggested that the protein for the 130-kDa glycoprotein receptor was encoded by clone a, and that the same protein also served as a core protein for the approximately 150-kDa heparan sulfate-proteoglycan receptor. Heparan sulfate glycosaminoglycan attachment to the 150-kDa receptor encoded by clone b was not detectable by the same enzyme treatment. Site-directed mutagenesis (from Ser to Ala) studies of the consensus sequence for the attachment of glycosaminoglycans further supported the presence of covalently attached heparan sulfate glycosaminoglycan in the approximately 150-kDa heparan sulfate-proteoglycan receptor. These receptors overexpressed in PT-r cells changed ligand accessibility or apparently translocated after changing extracellular calcium concentrations in a manner similar to the native receptors in PT-r cells (Sakaguchi, K. (1992) J. Biol. Chem. 267, 24554-24562), whereas those expressed in CHO-K1 or NIH/3T3 cells did not. These findings strongly suggest that the two FGF receptor isoforms cloned here represent the acidic FGF receptors that we reported earlier. A subpopulation of the receptors carries heparan sulfate glycosaminoglycan covalently attached to the core protein, and the change in ligand accessibility in response to the shift in ambient calcium concentration is specific to the parathyroid cells.

Successful allotransplantation of microencapsulated parathyroids in rats.

Allotransplantation of parathyroid tissue in humans is desirable for treating long-term hypoparathyroidism (e.g., after inadvertent removal of parathyroid glands during thyroid surgery). Until now, parathyroid allotransplantation was not used clinically because its advantages were outweighted by the need of immunosuppression. To overcome the immunogenicity of the tissue to be transplanted, we employed the method of microencapsulation; first tried in islet cell transplantation for experimental allotransplantation of parathyroid tissue. We have been able to achieve long-term success in a rat model. After isolation and tissue culture, tissue pieces from parathyroid glands of 280 Lewis rats were encapsulated in barium alginate and grafted into hypocalcemic DA rats. From the 7th to the 90th day after transplantation the recipient rats (DA rats) showed a normal serum calcium concentration. This is the first report of successful long-term survival and function of microencapsulated allotransplanted parathyroid tissue.

Cloning of a Parathyroid Hormone/Parathyroid Hormone-Related Peptide Receptor (PTHR) cDNA from a Rat Osteosarcoma (UMR 106) Cell Line: Chromosomal Assignment of the Gene in the Human, Mouse, and Rat Genomes

Complementary DNAs spanning the entire coding region of the rat parathyroid hormone/parathyroid hormone-related peptide receptor (PTHR) were isolated from a rat osteosarcoma (UMR 106) cell-line cDNA library. The longest of these clones (rPTHrec4) was used to chromosomally assign the PTHR gene in the human, rat, and mouse genomes. By somatic cell hybrid analysis, the gene was localized to human chromosome 3 and rat chromosome 8; by in situ hybridization, the gene was mapped to human chromosome 3p21.1-p22 and to mouse chromosome 9 band F; and by interspecific backcross analysis, the Pthr gene segregated with the transferrin (Trf) gene in chromosome 9 band F. Mouse chromosome 9 and rat chromosome 8 are known to be highly homologous and to also show synteny conservation with human chromosome 3. These three chromosomes share the transferrin gene (TF), the myosin light polypeptide 3 gene (MYL3), and the acylpeptide hydrolase gene (APEH). Our results add a fourth gene, the PTHR gene, to the synteny group conserved in these chromosomes.

Determination of bioactive rat parathyroid hormone (PTH) concentrations in vivo and in vitro by a 2-site homologous immunoradiometric assay

A new homologous 2-site assay for rat parathyroid hormone (IRMA), developed by Immutopics, Inc., has been evaluated and compared with a bone cell cAMP bioassay. Circulating PTH for adult rats assayed with this IRMA are in the range 10–15 pg/ml, and of the same order of magnitude as published values for biologically active PTH. The standard curve for the IRMA was linear over the range 3.4–240 pg/ml of rPTH 1–34, and serum samples diluted in parallel with the standard curve. The within-assay and between-assay coefficients of variation ranged from 5.2% ( n=18) to 7.6% ( n=24) and 8.3% ( n=16) to 26.4% ( n=10), respectively. Serum PTH values (mean±S.E.) for parathyroidectomized rats were 3.5±0.6 pg/ml ( n=18) versus 10.3±1.4 pg/ml ( n=16) for intact non-mated rats. Calcium injections suppressed circulating PTH by 50%. Lactating rats had serum PTH levels 5-fold higher and vitamin D deficient rats 60-fold higher than non-mated controls. PTH secreted from parathyroid cells in vitro was in the range 60–490 pg/ml as determined by the IRMA. These values represented 86.0±9.0% of the comparable bioassay values, indicating that the IRMA detects only bioactive PTH.

Establishment of a rapid bone resorption in vitro assay using previiously frozen mouse unfractionated bone cells pretreated with parathyroid hormone

We established a useful assay system for evaluating osteoclast-mediated bone resorption based on the use of unfractionated bone cells obtained from 10- to 11-day-old mice. When cells from 10 to 11 mice were treated for 7 days with rat parathyroid hormone (rPTH, 10 −8 M), a total of 4 to 5 × 10 7 cells could be obtained from the culture by treatment with 0.05% trypsin and 0.02% EDTA in PBS. These harvested cells contained about 20% tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells. When the harvested cells were cultured on dentine slices without rPTH, after 1 day, they formed TRAP-positive multinucleate cells that were active in bone rsorption. Eel calcitonin (eCT) decreased the number of pits in a dose-dependent manner, and its half maximal inhibition dose (ID 50) was 1.08 × 10 −11 M. Even after having been frozen in liquid nitrogen for 5 months, upon thawing, these cells were capable of forming pits; and this pit formation was inhibited by eCT. Since no appropriate osteoclaslic cell line for evaluating bone resorption is available at present, this system can provide a useful, practical means for assaying osteoclastic bone-resorbing activity.

Cysteine proteinases in rat parathyroid cells with special reference to their correlation with parathyroid hormone (PTH) in storage granules.

To further understand the roles of storage granules in parathyroid cells, we examined by immunocytochemistry the localization of cathepsins B and H and of PTH in rat parathyroid gland. In semi-thin sections, small and large granular immunodeposits for cathepsins B and H appeared in the cells, whereas those for PTH were detected throughout the cells, especially in perinuclear regions. By electron microscopy, immunogold particles indicating cathepsins B and H labeled lysosomes and storage granules, whereas those showing PTH were localized in storage granules, small secretory granules, and the trans-Golgi network. Small vesicles labeled by immunogold particles showing these proteinases often appeared close to the storage granules. By double immunostaining, immunogold particles indicating these proteinases were co-localized with those for PTH in storage granules. By EDTA treatment, immunoreactivity for cathepsins B and H and for PTH was notably reduced in the cells, but immunoreactivity for the proteinases was still seen in lysosomes. These results suggest that storage granules in the rat parathyroid cells fuse with small vesicles containing cathepsins B and H, which may participate in regulating the intracellular PTH levels by degrading PTH in the granules.

Stimulatory action of parathyroid hormone on renin secretion in vitro: a study using isolated rat kidney, isolated rabbit glomeruli and superfused dispersed rat juxtaglomerular cells.

1. We have previously reported that pharmacological concentrations (125nmol/l) of parathyroid hormone may stimulate renin release in the stable recirculating and non-filtering isolated rat kidney. 2. In the present study we have attempted to extend these initial observations by examining the concentration-related response of renin release to parathyroid hormone, using the same model of isolated kidney, and determining whether the effect of parathyroid hormone on renin release can be demonstrated by more direct approaches. Thus, the effects of parathyroid hormone on renin secretion were investigated in two other renal preparations: isolated rabbit glomeruli and isolated rat juxtaglomerular cells. 3. In the isolated kidney, rat parathyroid hormone significantly stimulated renin accumulation in the perfusate in a concentration-related manner with a threshold of 1 nmol/l. 4. In both glomeruli and juxtaglomerular cells bovine [Nle8,18,Tyr34]parathyroid hormone-(1-34)amide effectively and repeatedly stimulated renin release. These results imply that there is a direct stimulatory effect of parathyroid hormone on renin release. 5. We also examined the effect of [Nle8,18,Tyr34]parathyroid hormone-(1-34)amide during extracellular calcium buffering in the glomeruli. [Nle8,18,Tyr34]parathyroid hormone-(1-34)amide was uneffective in calcium-free medium. Increasing the extracellular ionized calcium concentration to 2.5 mmol/l increased the extent of stimulation in accordance with the reported ability of parathyroid hormone to block calcium channels and relax vascular smooth muscle cells. 6. These results provide further support for the role of parathyroid hormone as a direct mediator of renin secretion; moreover, the renin-stimulating action of parathyroid hormone may be mediated through the inhibition of calcium influx.

Estrogen receptors and biologic response in rat parathyroid tissue and C cells.

The expression of the PTH and calcitonin genes is dramatically decreased by 1,25(OH)2D3 in vivo, and the PTH gene expression is increased by hypocalcemia. We have now studied the effect of estrogens on the expression of these genes in vivo. 17 beta-Estradiol, given to ovariectomized rats, led to a fourfold increase in PTH mRNA and calcitonin mRNA levels. These effects occurred 24 h after single injections of 37-145 nmol estradiol, or after constant infusions of 12 pmol/d for 1 or 2 wk, where there was no effect on serum calcium levels. The estrogen receptor mRNA was demonstrated in the thyroparathyroid tissue by polymerase chain reaction. The estrogen binding was localized to the parathyroid and C cells by immunohistochemistry. Uterus weight was increased by repeated larger doses (73 nmol/d x 7) of estradiol, but not by the small doses (12 pmol/d for 1 or 2 wk) which were effective on the PTH and calcitonin genes, suggesting a sensitive endocrine effect. These results confirm that the parathyroid and C cells are target organs for estrogen, leading to an increased expression of PTH and calcitonin, which by their combined anabolic effect on bone would help prevent osteoporosis.