Effect of parathyroid hormone and cyclic adenosine 3',5'-monophosphate on isotonic fluid reabsorption: polarity of proximal tubular cells.

Isotomic fluid reabsorption (JV) of rat renal proximal tubules was examined by the shrinking droplet method in combination with simulatneous perfusion of blood capillaries. Sensitivity of JV measurement was improved by using each punctured tubule for control measurements: 1) Parathyroid hormoen (PTH) on the contraluminal cell side reduced JV in a dose-response behavior. The maximal inhibition was achieved at a PTH concentration of 10(-5) M, the half maximal inhibition at a concentration of 3 X 10(-9) M. PTH on the luminal cell side had a small inhibitory effect. 2) Cyclic AMP inhibited JV preferentially when applied to the luminal cell side. On the luminal cell side, both cyclic AMP and dibutyryl cyclic AMP inhibited JV in a similar dose-dependent behavior. Concentrations of both nucleotides as low as 10(-10) M had a definite inhibitory effect. Tested at a high concentration, N6-butyryl cyclic AMP was almost as effective as cyclic AMP. Deoxy cyclic AMP, 5' AMP, cyclic guanosine monophosphate (cyclic GMP), dibutyryl cyclic GMP had no effect. ATP inhibited JV to a very small extent. 3) The reduction of JV after administration of PTH and dibutyryl cyclic AMP was not additive. The similar inhibitory effect of PTH at the contraluminal cell face and of cyclic AMP at the luminal cell face suggests the following sequence of events in the mediation of the action of PTH: 1) activation of adenylate cyclase by PTH in the contraluminal cell membrane, and 2) action of the generated cyclic AMP on the luminal cell membrane. The interaction of cyclic AMP and the luminal cell membrane is initiated at the luminal cell surface.     

Effects of cyclic versus daily hPTH(1-34) regimens on bone strength in association with BMD, biochemical markers, and bone structure in mice.

We developed a cyclic PTH regimen with repeated cycles of 1-week on and off daily PTH injection and explored its effects on bone strength, BMD, bone markers, and bone structure in mice. Cyclic protocols produced 60-85% of the effects achieved by daily protocols with 57% of the total PTH given, indicating more economic use of PTH. The study supports further exploration of cyclic PTH regimens for the treatment of osteoporosis. INTRODUCTION: To minimize the cost and the catabolic action of hPTH(1-34), a cyclic PTH regimen with repeated 3-month cycles of on-and-off daily injection of hPTH(1-34) was developed in humans and shown to be as effective as a daily regimen in increasing vertebral BMD. However, changes in BMD may not adequately predict changes in bone strength. A murine model was developed to explore the efficacy of a cyclic PTH regimen on bone strength in association with other bone variables. MATERIALS AND METHODS: Twenty-week-old, intact, female C57BL/J6 mice (n = 7/group) were treated with (1) daily injection with vehicle for 7 weeks (control); (2) daily injection with hPTH(1-34) (40 microg/kg/day) for 7 weeks (daily PTH); and (3) daily injection with hPTH(1-34) and vehicle alternating weekly for 7 weeks (cyclic PTH). BMD was measured weekly by DXA, and serum bone markers, bone structure, and strength were measured at 7 weeks. RESULTS: Daily and cyclic PTH regimens increased BMD at all sites by 16-17% and 9-12%, respectively (all p < 0.01). The most dramatic effect of cyclic PTH occurred during the second week of treatment when PTH was off, with femoral and tibial BMD continuing to increase to the same extent as that produced by daily PTH. Both daily and cyclic PTH regimens significantly increased osteocalcin (daily, 330%; cyclic, 260%), mTRACP (daily, 145%; cyclic, 70%), femoral cortical width (daily, 23%; cyclic, 13%), periosteal circumference (daily, 5%; cyclic, 3.5%), and bone strength (max load: daily, 48%; cyclic, 28%; energy absorbed: daily, 103%; cyclic, 61%), respectively. Femoral bone strength was positively correlated with BMD, bone markers, and cortical structure. Neither regimen had an effect on vertebral bone strength. Although actual effects of cyclic PTH were 60-85% of those produced by daily PTH, the effects of cyclic PTH per unit amount administered were slightly greater than those of daily PTH for most measures. CONCLUSIONS: PTH-enhanced femoral bone strength is positively correlated with its effects on femoral BMD, bone markers, and bone structure. Cyclic PTH regimens represent a potential economic use of PTH and warrant further study.     

Characteristics of parathyroid hormone-specific cyclic changes of glucose-6-phosphate dehydrogenase activity in the distal convoluted tubule of the guinea pig

The effect of parathyroid hormone (PTH) on the time course of glucose-6-phosphate dehydrogenase (G6PD) activity in the distal convoluted tubule of a vitamin D-depleted guinea pig was determined using quantitative cytochemistry. G6PD activity decreased to the stable basal level 5 hrs after the initiation of the kidney segment maintenance cultures. The exposure of the tissues to 1 pg/ml of bovine PTH-(1-84) induced a cyclic change of G6PD activity, whereas neither carboxyl-terminal PTH nor other hormones tested showed such activity. After a 16-min exposure to bovine PTH-(1-84), the peak height of each cycle began to decrease until it disappeared at 34 min. The second exposure to this hormone at 46 min reinduced a similar cyclic change with a similar peak, indicating full viability of the cells. When bovine PTH-(1-84) was incubated with an excess amount of anti-bovine PTH antibody, the PTH-induced G6PD activity was completely abolished. Throughout a 14-min exposure to either human PTH-(1-84), human PTH-(1-34) or bovine PTH-(1-84), similar cyclic changes were observed with the constant peak height regardless of the dose (10(-16)-10(-12) M), although the cycle length shortened progressively as the dose was increased. They were equipotent on a molar basis between the concentrations of 10(-16) and 10(-13) M at 6 min of hormone exposure. The present data demonstrate that the cytochemical bioassay of PTH in a vitamin D-depleted animal is based on a dose-dependent difference in the time course of G6PD activity.     

Mechanism of dopamine inhibition of renal phosphate transport.

Dopamine (DA) is natriuretic and phosphaturic. However, whether the effect of DA on Pi reabsorption is a consequence of its effect on sodium transport is not known. Therefore, this study was performed to determine the effect of DA on the maximal transport of phosphate (TmPi), and upon the capacity of renal proximal brush border membrane (BBM) for (Naextra-vesicular greater than Naintravesicular)-gradient-dependent transport of Pi, as compared with the transport of other solutes. Graded infusions of Pi (0, 1, 2, and 3 mumols/min) were given to thyroparathyroidectomized male Sprague-Dawley rats in the presence of vehicle (0.9% NaCl; N = 5), DA 15 micrograms/kg/min; N = 6), or parathyroid hormone ((PTH); 1 U/kg/min; N = 5). The TmPi for rats infused with DA (3.3 +/- 0.3 mumol/mL) was significantly less than the TmPi for saline control rats (4.4 +/- 0.2 mumol/mL). Rats infused with PTH exhibited the lowest TmPi (1.8 +/- 0.3 mumol/mL). No differences in sodium excretion were observed among any of the groups. Na-dependent Pi transport was studied in BBM vesicles (BBMV) prepared from rats fed a low-phosphate diet for 2 days that were anesthetized, acutely thyroparathyroidectomized, and systemically infused with DA (350 micrograms bolus, plus 35 micrograms/kg/min; N = 8), PTH (33 U/kg bolus, followed by a continuous infusion of 1 U/kg/min; N = 6), or vehicle (1 mL/kg bolus, plus 2 mL/h constant infusion of 0.9% NaCl; N = 8) for 90 min. DA significantly inhibited the Na cotransport of Pi by 22.4 +/- 4.1% (P less than 0.01) as compared with the control group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Responses of Urinary N-Telopeptide and Renal Calcium Handling to PTH Infusion after Treatment with Estrogen, Raloxifene, and Tamoxifen

This prospective, randomized, placebo-controlled study investigated whether estrogen, tamoxifen, and raloxifene protect the skeleton from the acute catabolic effects of continuous PTH(1–34) infusion. It was infused over 24 h in 25 postmenopausal women both before and while on medication for 16–20 weeks (estrogen n = 7, raloxifene n = 5, tamoxifen n = 7, placebo n = 6). Blood and urine samples were collected at baseline and every 4 h during the PTH(1–34) infusion and analyzed for calcium homeostasis, bone remodeling, and specific cytokines. Data for the premedication PTH(1–34) infusions were pooled. During the premedication PTH(1–34) infusions, serum calcium and urine phosphorus increased, while serum phosphorus and urine calcium declined. Osteocalcin decreased (mean 18%), while urine NTX increased (mean 315%). Serum IL-6 increased 260%, but there were no other cytokine changes as a result of the PTH(1–34) infusion. On medication, the mean peak change in NTX with PTH(1–34) infusion was less (77, 59, and 31 nM/mM with raloxifene, tamoxifen, and estrogen, respectively). The reduction in urine calcium excretion was prolonged with each agent but only significantly with estrogen. There was no reduction in the IL-6 elevation induced by PTH(1–34) with any medication. The differential skeletal resorption response to PTH(1–34) infusion after the treatments may reflect different potencies of these agents or variability in interaction with the estrogen receptor. Renal calcium conservation and the blunted response of bone resorption to PTH(1–34) infusion may be mechanisms by which estrogen and estrogen agonist/antagonist agents preserve bone mass.     

Calcemic and phosphaturic response to parathyroid hormone in normal and chronically uremic dogs

It is well established that the calcemic response to parathyroid hormone (PTH) is blunted in chronic uremia and is corrected partially by 1,25(OH)2D treatment. Recent evidence suggests that PTH(1-34) and not PTH(1-84) may be the actual calcemic fragment. Equivalent doses of both peptides were infused into five normal dogs (GFR = 51 ml/min) and eight dogs with a remnant kidney and chronic renal insufficiency (GFR = 15 ml/min). Both the calcemic and phosphaturic responses were studied. Remnant dogs had a blunted calcemic response to bPTH(1-84). The increase in fractional phosphate excretion was similar. In contrast, the calcemic response to bPTH(1-34) was equivalent in remnant and normal dogs. Treatment of uremic dogs with 400 ng 1,25(OH)2D daily for 3 days restored the effect of bPTH(1-84) on serum calcium and increased the control value for tubular phosphate reabsorption from 28 +/- 3 micrograms/ml GFR to 37 +/- 4 micrograms/ml GFR (P less than 0.01). These results suggest that there is an impaired conversion of PTH(1-84) to PTH(1-34) in chronic renal insufficiency and that 1,25 (OH)2D may be involved in metabolism of PTH(1-84). In addition, the effect of PTH on fractional phosphate excretion is not magnified in nonparathyroidectomized uremic dogs.     

Pacing-induced cardiac failure of hypertrophic hearts: effects of cyclic GMP reduction.

BACKGROUND: We tested the hypothesis that pacing-induced cardiac failure of hypertrophic hearts would reduce the functional and metabolic responses of these hearts to guanylate cyclase inhibition and this was associated with alterations in cyclic GMP. MATERIALS AND METHODS: Methylene blue (MB, 2 mg/kg/min, guanylate cyclase inhibitor) was infused into the left anterior descending coronary artery in 5 control, 5 left ventricular hypertrophy (LVH), and 5 LVH pacing-induced failure dogs. Regional myocardial work was calculated as the integrated product of force and segment shortening and regional myocardial O(2) consumption (VO(2)) from coronary blood flow and O(2) extraction measurements. Cyclic GMP was determined by radioimmunoassay. RESULTS: MB increased regional work (635 +/- 169 vs 1649 +/- 500, 781 +/- 184 vs 1569 +/- 203 g * mm/min) and VO(2) (8.3 +/- 1.4 vs 10.9 +/- 1.4, 7.3 +/- 0.7 vs 9.1 +/- 0.7 ml O(2)/min/100 g) in both control and LVH dogs but not in failure dogs (536 +/- 234 vs 623 +/- 193, 3.6 +/- 1.1 vs 4.7 +/- 1.9). MB also decreased cyclic GMP in control dogs (1170 +/- 142 vs 812 +/- 105 pmol/g). LVH dogs had elevated baseline cyclic GMP (5875 +/- 949) compared to control dogs but also demonstrated decreased cyclic GMP in response to MB (2820 +/- 372). In failure dogs, basal cyclic GMP was also elevated (4650 +/- 613) compared to control dogs but there was a lack of response to MB (3670 +/- 640). CONCLUSIONS: We conclude that the myocardial function, VO(2) and cyclic GMP responses to methylene blue are diminished in the transition from hypertrophy to cardiac failure.     

Hormonal and nonhormonal desensitization in isolated bone cells

Summary Prior exposure to PTH markedly decreased the responsiveness of isolated, cultured bone cells to the stimulatory effect of the hormone on cyclic AMP formation. This process of desensitization developed within 30 min, persisted during prolonged incubation of the cells in PTH-free medium, and could not be attributed to enhanced excretion of cyclic AMP from the cells, nor to the extracellular accumulation of an inhibitor of PTH action. Adenylate cyclase activity in a subcellular fraction derived from PTH-treated cells was refractory to PTH and to sodium fluoride. These results indicate that PTH-mediated desensitization reflects, at least in part, impaired cyclic AMP formation. Adenosine and PGE₂, known stimulators of bone cell cyclic AMP formation, elicited agonist-specific desensitization, and also desensitized bone cells to the effects of subsequently added PTH. PTH blunted the cellular response to adenosine, but not to PGE₂. Modest refractoriness to PTH was evident in cells that had been treated previously with the cyclic AMP phosphodiesterase inhibitors IBMX, theophylline, and Bt₂cAMP, whereas treatment with sodium butyrate had no effect. The actions of the inhibitors, like that of PTH, were rapid in onset and long-lasting. Desensitization caused by previous treatment with the phosphodiesterase inhibitors, and with PTH itself, was accompanied by enhanced phosphodiesterase activity in bone cell homogenates. Induction of phosphodiesterase activity may well contribute to desensitization in the bone cell system. Abbreviations used in this paper: HBSS, Hank's balanced salt solution; EBSS, Earle's balanced salt solution; IBMX, 3 isobutyl 1 methyl xanthine; PTH, synthetic bovine parathyroid hormone (amino acids 1-34); PGE₁, prostaglandin E₁; PGE₂, prostaglandin E₂; Bt₂cAMP, N⁶,O²′ dibutyryl cyclic 3′,5′ adenosine monophosphate.     

The PTH-Calcium Relationship During a Range of Infused PTH Doses in the Parathyroidectomized Rat

To establish the PTH dosage that maintains normal mineral homeostasis in the PTX rat, a series of doses of rat 1-34 PTH were infused via a subcutaneously implanted miniosmotic pump. The doses were 0, 0.011, 0.022, 0.044, and 0.11 μg/100 g/hour. After 48 hours, serum calcium ranged from 5.56 ± 0.02 to 16.29 ± 0.25 mg/dl, ANOVA P < 0.001, and serum phosphorus from 12.49 ± 0.03 to 5.33 ± 0.34 mg/dl, ANOVA P < 0.001. By post hoc test, the serum calcium level was different (P < 0.05) at every PTH dose; the serum phosphorus level was different (P < 0.05) at every PTH dose except between the two highest doses. The PTH dosage that produced a normal serum calcium (10.09 ± 0.10 mg/dl) and phosphorus (6.90 ± 0.18 mg/dl) was 0.022 μg/100 g/hour. The relationship between increasing doses of PTH and both serum calcium and phosphorus was curvilinear and the calcium-phosphorus product was remarkably constant from a serum calcium of 7–13 mg/dl. The increase in serum calcium and the decrease in serum phosphorus were more rapid at lower than at higher PTH doses so that for both, an asymptote was reached. At the highest serum calcium values, the calcium-phosphorus product increased and in individual rats, an increase in serum phosphorus was associated with a decrease in serum calcium. In summary, this study shows that (1) for rat 1-34 PTH, the normal replacement dose in the PTX rat with normal renal function on a normal diet is 0.022 μg/100 g/hour; (2) the relationship between PTH and both serum calcium and phosphorus is curvilinear, and an asymptote is reached for both; and (3) the calcium-phosphorus product is remarkably constant as the serum calcium increases from 7 to 13 mg/dl and only increased during marked hypercalcemia when serum phosphorus did not decrease further or even tended to increase. Received: 30 May 1997 / Accepted: 15 October 1997     

Parathyroid hormone stimulation of mitosis in rat thymic lymphocytes is independent of cyclic AMP

The in vitro mitotic response of rat thymic lymphocytes to hPTH(1-34), hPTH (1-38), and 8,18 Nle hPTH(1-34) exhibits a dependency upon extracellular calcium. Removal of extracellular calcium or the addition of Verapamil (5 micrograms/ml) or trifluoroperazine (10 microM) abrogated the mitotic response. Mitogenic concentrations of 8,18 Nle hPTH(1-34) increased calcium 45 uptake from 4.49 +/- 0.25 to 8.23 +/- 0.75 pMol/10(6) cells/min. The intracellular calcium concentration, measured by Quin 2 fluorescence, also increased after addition of 8,18 Nle hPTH(1-34). Parathyroid hormone-induced activation could not be demonstrated in an otherwise responsive thymocyte membrane adenylate cyclase. In intact cells mitogenic levels of 8,18 Nle hPTH(1-34) decreased intracellular cyclic AMP content. This response was blocked by both 3-isobutyl 1-methyl xanthine and trifluoroperazine, and may indicate activation of calcium-dependent phosphodiesterase. We conclude that PTH stimulates thymic lymphocyte proliferation independently of cyclic AMP, and that changes in cellular calcium homeostasis are intimately involved in the action of PTH. In all of the assays employed, the hitherto antagonistic analogue 8,18 Nle 34 Tyr bPTH(3-34)amide proved to be an agonist. We postulate that the receptor utilized for this PTH action may not exhibit classical PTH structure-activity specificities.     

Alkaline phosphatase inhibition by parathyroid hormone and isoproterenol in a clonal rat osteosarcoma cell line. Possible mediation by cyclic AMP

Summary The effect of parathyroid hormone (PTH 1–34 bovine) on alkaline phosphatase activity was investigated in an osteoblast-like clonal cell line derived from rat osteosarcoma (ROS 17/2). ROS 17/2 alkaline phosphatase resembled the bone enzyme in levamisole sensitivity and electrophoretic mobility but differed in heat stability. The specific activity of ROS 17/2 alkaline phosphatase increased with time in culture. This increase was inhibited by PTH (1–34) and (-)-isoproterenol in a dose-dependent manner starting at near-physiological hormone concentrations. The ID₅₀ values were 0.02 nM for PTH (1–34) and 1.7 nM for isoproterenol. The two hormones stimulated ROS 17/2 adenylate cyclase, albeit at higher concentrations: K_m values were 13 nM for PTH (1–34) and 16 nM for isoproterenol. The rise in alkaline phosphatase was also inhibited by dibutyryl cyclic AMP and 8-bromocyclic AMP (0.1 mM). These findings further document the osteoblastic properties of the ROS 17/2 osteosarcoma cell line, suggest that PTH inhibition of alkaline phosphatase represents a physiological response to the hormone in these cells, and implicate cyclic AMP as a mediator of this PTH effect.     

Abaloparatide at the Same Dose Has the Same Effects on Bone as PTH (1-34) in Mice

Abaloparatide, a novel analog of parathyroid hormone-related protein (PTHrP 1–34), became in 2017 the second osteoanabolic therapy for the treatment of osteoporosis. This study aims to compare the effects of PTH (1-34), PTHrP (1-36), and abaloparatide on bone remodeling in male mice. Intermittent daily subcutaneous injections of 80 μg/kg/d were administered to 4-month-old C57Bl/6J male mice for 6 weeks. During treatment, mice were followed by DXA-Piximus to assess changes in bone mineral density (BMD) in the whole body, femur, and tibia. At either 4 or 18 hours after the final injection, femurs were harvested for μCT analyses and histomorphometry, sera were assayed for bone turnover marker levels, and tibias were separated into cortical, trabecular, and bone marrow fractions for gene expression analyses. Our results showed that, compared with PTH (1-34), abaloparatide resulted in a similar increase in BMD at all sites, whereas no changes were found with PTHrP (1-36). With both PTH (1-34) and abaloparatide, μCT and histomorphometry analyses revealed similar increases in bone volume associated with an increased trabecular thickness, in bone formation rate as shown by P1NP serum level and in vivo double labeling, and in bone resorption as shown by CTX levels and osteoclast number. Gene expression analyses of trabecular and cortical bone showed that PTH (1-34) and abaloparatide led to different actions in osteoblast differentiation and activity, with increased Runx2, Col1A1, Alpl, Bsp, Ocn, Sost, Rankl/Opg, and c-fos at different time points. Abaloparatide seems to generate a faster response on osteoblastic gene expression than PTH (1-34). Taken together, abaloparatide at the same dose is as effective as PTH (1-34) as an osteoanabolic, with an increase in bone formation but also an increase in bone resorption in male mice. © 2019 American Society for Bone and Mineral Research.     

Selective phosphodiesterase 5 inhibition does not reduce propofol sedation requirements but affects speed of recovery and plasma cyclic guanosine 3',5'-monophosphate concentrations in healthy volunteers

Cyclic guanosine 3',5'-monophosphate (cyclic GMP) has been implicated in modulating the effects of anesthesia. We hypothesized that limiting the breakdown of cyclic GMP through selective phosphodiesterase inhibition would influence propofol sedation requirements and plasma cyclic GMP concentrations. Ten volunteers received 100 mg of sildenafil or placebo orally in this placebo-controlled, double-blind, randomized crossover pilot study. Propofol sedation was achieved using a target-controlled infusion system until loss of verbal contact (LVC). Plasma cyclic GMP concentrations were determined at baseline, LVC, and 30 min after LVC. There was no difference in the amount of propofol used, predicted plasma concentration, or duration of sedation in volunteers after sildenafil compared with placebo treatment. Return of spontaneous verbal contact was faster after sildenafil (4 [3-8] min versus 6 [3-5] min, median [range], P = 0.019). Cyclic GMP concentrations were reduced during propofol sedation in the placebo group compared with baseline (P < 0.004). The plasma cyclic GMP concentrations were larger (P = 0.004) at LVC in the sildenafil group compared with placebo. We have shown that selective phosphodiesterase 5 inhibition decreases recovery time from propofol sedation without affecting propofol requirements. The decrease of plasma cyclic GMP concentrations during propofol sedation in the placebo group indicates a potential role of cyclic GMP in propofol anesthesia in humans. IMPLICATIONS: Plasma cyclic guanosine 3',5'-monophosphate (cyclic GMP) concentrations are reduced during propofol sedation. Selective phosphodiesterase 5 inhibition, however, does not reduce propofol sedation requirements or plasma cyclic GMP concentrations but affects speed of recovery in healthy volunteers.     

Atrial natriuretic peptide and parathyroid hormone (1-84) in relation to noradrenaline induced changes in blood pressure in uraemic and healthy subjects.

In order to evaluate the hormonal regulation of blood pressure (BP) in uraemia 12 patients on chronic maintenance dialysis and 14 healthy controls were studied. BP and plasma concentrations of atrial natriuretic peptide (ANP), cyclic 3',5'-guanosine monophosphate (cGMP), and intact parathyroid hormone (PTH(1-84)) were determined before, during, and after a 60 min noradrenaline infusion 0.1 micrograms kg-1 body wt. min-1. Mean BP increased to the same extent in the uraemic patients (median 15 mmHg, range 6-25 mmHg) as in the controls (12 mmHg, 5-25 mmHg). ANP increased during noradrenaline infusion both in patients (7.2 to 8.3 pmol/l, medians, p < 0.01) and in controls (4.4 to 6.0 pmol/l, p < 0.01), and so did cGMP (patients: 31.6 to 35.9 nmol/l, p < 0.05; controls: 6.6 to 8.7 nmol/l, p < 0.01). PTH(1-84) was higher in the uraemic patients than in the controls, but was unchanged during noradrenaline infusion in both groups. Correlation analyses gave no evidence of a direct relation between BP and ANP, but basal PTH(1-84) was negatively correlated to basal mean BP in the patients (rho = -0.615, p < 0.05), but not in the controls. In conclusion, noradrenaline induced similar elevations of BP in dialysis patients as in healthy controls despite elevated ANP and PTH(1-84) in the patients, and ANP release was stimulated in both groups. PTH(1-84) may participate in blood pressure regulation in uraemic patients.     

Effects of cyclic vs. daily treatment with human parathyroid hormone (1-34) on murine bone structure and cellular activity

Previously, we demonstrated that the human parathyroid hormone (1-34) fragment (hPTH(1-34)) increased bone strength in proportion to its effects on BMD and cortical bone structure in the murine femur by comparing cyclic vs. daily administration of hPTH(1-34). Both cyclic and daily regimens increased vertebral BMD similarly at 7 weeks. Here, we have examined the effects of daily and cyclic PTH regimens on bone structure and cellular activity by static and dynamic histomorphometry. Twenty-week-old, intact female C57BL/J6 mice were treated with the following regimens (n=7 for each group): daily injection with vehicle for 7 weeks [control]; daily injection with hPTH(1-34) (40 microg/kg/day) for 7 weeks [daily PTH]; and daily injection with hPTH(1-34) (40 microg/kg/day) and vehicle alternating weekly for 7 weeks [cyclic PTH]. At days 9 and 10, and 2 and 3 prior to euthanasia, calcein (10 mg/kg) was injected subcutaneously. At the end of study, the lumbar vertebrae 1-3 and the left femora were excised, cleaned, and processed for histomorphometry. In the lumbar vertebrae, daily and cyclic PTH regimens significantly increased cancellous bone volume (BV/TV), trabecular number, trabecular osteoclast and osteoblast perimeters, trabecular mineral apposition rate (MAR) and bone formation rate (BFR), and periosteal MAR and BFR compared to control, with no significant difference between the two PTH-treated groups. Increased trabecular tunneling was observed in both PTH-treated groups. Both regimens tended to increase vertebral cortical bone formation parameters with the effects at the periosteum site being more marked than those at the endosteum site, resulting in a significant increase in cortical width. In the femur, the effects of cyclic PTH on BV/TV, trabecular width and number, trabecular and endocortical osteoblast and osteoclast perimeters, cortical width, and trabecular and periosteal BFR were less marked than those of daily PTH. A cyclic PTH regimen was as effective as a daily regimen in improving cancellous and cortical bone microarchitecture and cellular activity in the murine vertebra.     

Comparison of bone formation responses to parathyroid hormone(1-34), (1-31), and (2-34) in mice

In this study we used a mouse model system to compare the in vivo effects of parathyroid hormone(1-34) [PTH(1-34)] with that of PTH(1-31) or PTH(2-34) analogs. Daily subcutaneous administration of PTH(1-34) for 15 days caused a dose-dependent increase in the serum osteocalcin level and bone extract alkaline phosphatase activity, markers of bone formation. PTH(2-34) was much less potent, whereas PTH(1-31) was equipotent in stimulating bone formation parameters in mice. PTH(1-34) caused significant increases in serum calcium (after 4 h) and tartrate-resistant acid phosphatase activity in bone extract (after 4 h), whereas PTH(2-34) and PTH(1-31) were less potent. Because PTH(1-31) caused a smaller increase in bone resorption parameters compared to PTH(1-34), despite similar effects on bone formation parameters, we evaluated the long-term anabolic effects of PTH(1-31) and PTH(1-34) in mice. Weekly evaluations of serum osteocalcin levels demonstrated that daily injections of PTH(1-34) and PTH(1-31) at 80 microg/kg body weight increased serum osteocalcin levels within 1 week of the start of treatment, which were maintained during the entire 22 week treatment. Assessment of bone density at the end of the treatment period with peripheral quantitated computed tomography (pQCT) revealed that PTH(1-34) caused a significantly greater increase in femoral bone density compared to PTH(1-31) at the middiaphysis (18% vs. 9% over vehicle control; p < 0.001). Both PTH(1-34) and PTH(1-31) increased periosteal circumference compared to vehicle (p < 0.01) without a significant difference between the two treatments. In contrast, PTH(1-34) caused a significantly greater reduction in endosteal circumference than PTH(1-31) (p < 0.001). Both analogs significantly increased maximum load and area of moment of inertia over the vehicle group. In conclusion, our findings suggest that PTH(1-34) and PTH(1-31) may exhibit different anabolic effects at the periosteum vs. endosteum in the long bones of mice.     

Relative sensitivity of kidney and bone to the amino-terminal fragment b-PTH (1–30) of native bovine parathyroid hormone: Implications for assessment of bioactivity of parathyroid hormone fragments in vivo and in vitro

Summary Adenylate cyclase activation in renal cortical membranes is widely used to assess the potency of parathyroid hormone (PTH) and hormonal fragments. Recent studies, however, suggest that the structural requirements for biological activity of PTH in bone may differ from those in kidney. In isolated perfused canine bone, cyclic AMP production is markedly increased by syn-b-PTH (1–34) whereas intact b-PTH (1–84) has minimal effect. Furthermore, a potent inhibitor of PTH stimulated adenylate cyclase activity, Nle⁸Nle¹⁸Tyr³⁴ syn-b-PTH (3–34) NH₂, devoid of biological activity in kidney, is an agonist in isolated bone. The present studies examine the effects in bone and kidney of the amino-terminal fragment b-PTH (1–30), prepared by cleavage of intact b-PTH (1–84) by Cathepsin Bin vitro. In basolateral canine renal cortical membranes, b-PTH (1–30) was less active than syn-b-PTH (1–34) in its ability to stimulate adenylate cyclase. Half maximal stimulation of adenylate cyclase activity by b-PTH (1–30) was 50 nM compared with 2 nM for syn-b-PTH (1–34). Maximum enzyme activation by b-PTH (1–30) reached only 50% of the activation by syn-b-PTH (1–34). Addition of Gpp(NH)p (1 mM) increased the affinities for both peptides. The relative difference in potency, however, remained unchanged. In contrast, in isolated bone, b-PTH (1–30) and syn-b-PTH (1–34) were equipotent in increasing cyclic AMP production. These data provide further evidence for differences in the structural requirements for PTH activity in bone and kidney and suggest that bioassays of PTH and PTH fragments in kidney may not accurately reflect the effects of PTH in bone.     

Effects of hPTH(1‐34) Infusion on Circulating Serum Phosphate, 1,25‐Dihydroxyvitamin D,and FGF23 Levels in Healthy Men

Fibroblast growth factor 23 (FGF23) promotes phosphaturia and suppresses 1,25‐dihydroxyvitamin D [1,25(OH)₂D] production. PTH also promotes phosphaturia, but, in contrast, stimulates 1,25(OH)₂D production. The relationship between FGF23 and PTH is unclear, and the acute effect of pharmacologically dosed PTH on FGF23 secretion is unknown. Twenty healthy men were infused with human PTH(1‐34) [hPTH(1‐34)] at 44 ng/kg/h for 24 h. Compared with baseline, FGF23, 1,25(OH)₂D, ionized calcium (iCa), and serum N‐telopeptide (NTX) increased significantly over the 18‐h hPTH(1‐34) infusion (p < 0.0001), whereas serum phosphate (PO₄) transiently increased and then returned to baseline. FGF23 increased from 35 ± 10 pg/ml at baseline to 53 ± 20 pg/ml at 18 h (p = 0.0002); 1,25(OH)₂D increased from 36 ± 16 pg/ml at baseline to 80 ± 33 pg/ml at 18 h (p < 0.0001); iCa increased from 1.23 ± 0.03 mM at baseline to 1.46 ± 0.05 mM at hour 18 (p < 0.0001); and NTX increased from 17 ± 4 nM BCE at baseline to 28 ± 8 nM BCE at peak (p < 0.0001). PO₄ was 3.3 ± 0.6 mg/dl at baseline, transiently rose to 3.7 ± 0.4 mg/dl at hour 6 (p = 0.016), and then returned to 3.4 ± 0.5 mg/dl at hour 12 (p = 0.651). hPTH(1‐34) infusion increases endogenous 1,25(OH)₂D and FGF23 within 18 h in healthy men. Whereas it is possible that the rise in PO₄ contributed to the observed increase in FGF23, the increase in 1,25(OH)₂D was more substantial and longer sustained than the change in serum phosphate. Given prior data that suggest that neither PTH nor calcium stimulate FGF23 secretion, these data support the assertion that 1,25(OH)₂D is a potent physiologic stimulator of FGF23 secretion.     

Beta1-adrenergic blockade augments pulsatile PTH secretion in humans

Pulsatile peptide hormone secretion provides efficient control of specific end organ functions. To test the hypothesis that sympathetic neuronal activity drives synchronous pulsatile PTH release from the parathyroid glands, we investigated the acute effects of beta1-adrenergic receptor blockade on PTH secretion patterns in a single-blinded study in nine healthy adults. Plasma PTH levels were determined at 1-min intervals. After a 75-min baseline period, seven subjects received a continuous intravenous infusion of the short-acting beta1-adrenergic receptor blocker esmolol for 105 min. After a 30-min washout period, esmolol was infused for another 30 min. Two additional subjects were randomized to receive solvent infusions. PTH secretion characteristics were analyzed by multiparameter deconvolution analysis, and the orderliness of plasma PTH fluctuations by Approximate Entropy statistics. BP slightly decreased during esmolol infusion, whereas heart rate, ionized calcium, phosphate, magnesium, and plasma and urine catecholamines remained unchanged. Esmolol increased mean plasma PTH by 33 +/- 8% (P < 0.01), due to a preferential increase in the pulsatile PTH secretion component (+129 +/- 44%, P < 0.02). The increased pulsatile PTH secretion was mediated by an augmented PTH burst mass (+117 +/- 42%, P < 0.01), whereas burst frequency remained unchanged. The regularity of PTH fluctuations was not affected by the beta-adrenergic blockade. The effects were reproducible during the second esmolol infusion. The authors conclude that the sympathetic nervous system has a modulating effect on pulsatile PTH secretion. Selective beta-1 adrenergic blockade acutely increases plasma PTH by augmenting the mass of hormone secreted per burst, but it does not alter the rhythmicity of pulsatile PTH release.