Howard Rasmussen

Protein kinase C in the regulation of smooth muscle contraction.

The cellular and molecular mechanisms underlying smooth muscle contraction are reviewed in the light of recent studies of smooth muscle ultrastructure and of the role of polyphosphoinositide turnover and protein kinase C function in smooth muscle contraction. A new model of smooth muscle contraction is proposed that differs radically from accepted views, particularly the latch bridge hypothesis, in terms of both Ca2+ messenger function and the molecular events underlying this process. A coordinate fibrillar domain model of contraction is proposed in which the initial and sustained phases of contraction are mediated by different cellular and molecular events. The initial phase of response is mediated by a rise in [Ca2+ ]c and the resulting calmodulin‐dependent activation of both myosin light chain kinase and the dissociation of caldesmon from the actin‐caldesmon‐tropomyosin‐myosin fibrillar domain. These events lead to an interaction between actin and the phosphorylated light chains of myosin just as in previous models. However, this initial phase is followed by a sustained phase in which a rise in [Ca2+ ]sm stimulates the plasma membrane‐associated, Ca2+ ‐sensitive form of protein kinase C that results in the phosphorylation of both structural and regulatory components of the filamin‐actin‐desmin fibrillar domain. These events underlie the tonic phase of contraction.— Rasmussen, H.; Takuwa, Y.; Park, S. Protein kinase C in the regulation of smooth muscle contraction. FASEB J. 1: 177‐185; 1987.

Hormonal control of the renal conversion of 25-hydroxycholecalciferol to 1,25-dihydroxycholecalciferol

Isolated renal tubules from vitamin D-deficient chicks catalyse the in vitro conversion of 25-hydroxycholecalciferol to 1,25-dihydroxycholecalciferol. This conversion is stimulated by 5 x 10(-10) M bovine parathyroid hormone, or by 10(-6) M cyclic AMP. It is inhibited by 10(-9) M porcine calcitonin. It is concluded that these hormonal controls of the synthesis of the renal hormone 1,25-dihydroxycholecalciferol are of particular physiological significance in coordinating the activities of the various organs involved in extracellular calcium homeostasis.

TPA-induced contraction of isolated rabbit vascular smooth muscle

Myosin light chain phosphorylation may not regulate the sustained phase of vascular smooth muscle contraction. Another, unidentified, calcium-dependent pathway may be involved in this process. TPA, an activator of C-kinase, at concentrations of 10 to 333 nM induces a calcium-dependent contraction of vascular smooth muscle which develops slowly but progressively to reach values of 50-300 mm Hg. Arteries exposed to the ionophore A23187, in a calcium-free medium, display a uniform series of contractile responses when exposed to 1.5 mM Ca2+ for 2 min once every 10 min. Exposure to 100 nM TPA as well as ionophore leads to a progressive enhancement of these calcium-induced, contractile responses. Arteries stimulated by brief (10 sec), repetitive (every 3 min) electrical pulses, respond with a series of comparable phase 1 responses. Prior exposure of vessels to 10 nM TPA, causes a progressive increase in the magnitude of these responses to repetitive electrical stimulation. Addition of 25 microM forskolin, an activator of adenylate cyclase, to TPA-treated, partially-contracted muscle leads to the immediate inhibition of the TPA-induced contraction. These data suggest that the activation of C-kinase plays a significant role in regulating vascular smooth muscle contraction.

Insulin secretion: Combined effects of phorbol ester and A23187

The effect of the ionophore, A23187, and/or the phorbol ester, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), on insulin secretion were compared with those of glucose. Glucose induces a biphasic pattern of insulin secretion; A23187 a comparable initial spike but no second phase; and TPA a slowly progressive increase. Combined A23187 and TPA evoke a pattern similar to that induced by glucose. Forskolin enhances both phases of glucose- induced and of TPA-A23187-induced insulin secretion. These results are interpreted in terms of a model of cell activation in which two branches of the calcium messenger system, the calmodulin branch and the C-kinase branch, control, respectively, the initial and sustained phases of insulin secretion.

The Importance of Circulating 1,25-Dihydroxyvitamin D in the Pathogenesis of Hypercalciuria and Renal-Stone Formation in Primary Hyperparathyroidism

Fifty patients with primary hyperparathyroidism were studied with an oral calcium-tolerance test, measurements of plasma levels of vitamin D metabolites, and determination of calcium excretion on both a low-normal (400 mg) and high-normal (1000 mg) calcium intake. There were strong positive correlations between plasma levels of 1,25-dihydroxyvitamin D (1,25(OH)2D) and both the calciuric response to the calcium-tolerance test (r = +0.75, P less than 0.001) and calcium excretion on the 1000-mg calcium diet (r = +0.65, P less than 0.001). The patients were classified into two subpopulations: 30 patients showed hyperabsorption with the calcium-tolerance test, striking hypercalciuria, marked elevations in plasma 1,25(OH)2D, and a high incidence (19 of 30 patients) of renal stones; 20 patients had a normal response to the tolerance test, normocalciuria, normal or high-normal plasma 1,25(OH)2D, and a low incidence of stones (three of 20 patients). The findings emphasize the importance of circulating 1,25(OH)2D in the pathogenesis of hypercalciuria and stone formation in primary hyperparathyroidism.

THE RELATIONSHIP BETWEEN VITAMIN D AND PARATHYROID HORMONE

Over three decades ago, it was established that deficiency of either vitamin D or parathyroid hormone may lead to tetany and hypocalcemia (1, 2). Since then, there has been continued interest in the relationship between the biologic effects of these two agents. At one time it was believed that the D vitamins exerted their effects by stimulating the parathyroid glands, but this concept became untenable after it was established that vitamin D could induce hypercalcemia in hypopara-thyroid organisms. More recently it has been proposed that the hormone exerts its characteristic effects only in vitamin D-fed animals (3), and that the basis of this action depends upon a chemical interaction between the two (4). These proposals are not satisfactory because there is considerable clinical and pathological evidence (5) suggesting that parathyroid hyperfunction exists in vitamin D deficiency. Certainly complete synergism of action seems most unlikely because of the well-established difference in the syndromes produced by their separate deficiencies (5). On the other hand, there is no satisfactory explanation for the persistent hypocalcemia in D-deficient animals, in spite of apparent parathyroid over-function, nor for the observations of Harrison, Harrison, and Park (3) that the administration of a standard dose of parathyroid hormone to a D-deficient rat results in little or no change in the concentration of either calcium or phosphate in the plasma. Another problem that has remained unresolved is the explanation of the phosphate retention that occurs in a D-deficient animal or human shortly after the initiation of specific therapy (6). Har-rison and Harrison (6) have favored the view that this retention is due to a direct action of vitamin D upon the renal tubular reabsorption of phosphate , whereas others (5) have considered it a consequence of decreased parathyroid gland activity. A possible new insight into the nature of the vitamin D and parathyroid hormone relationship has come from mitochondrial studies (7-11). It has been found that vitamin D and parathyroid hormone promote the release of calcium (as a phosphate salt or ion pair) from isolated mito-chondria (7), that they act synergistically, and that the presence of the vitamin is necessary for the hormone to exert this effect, although the converse is not true. The hormone has other effects upon the mitochondria that are neither produced by, nor dependent upon, the presence of vitamin D. These are the stimulation of phosphate (11), sulfate, and arsenate (9) accumulation, and a stimulation …

On the pathogenesis of so-called idiopathic hypercalciuria

Abstract Forty-seven patients were identified as having persistent hypercalciuria during a period of low calcium intake ( Type 2 patients were divided into two subtypes on the basis of their response to the administration of methylchlorothiazide and 25-hydroxyvitamin D 3 (25(OH)D 3 ). Type 2a patients responded to either treatment with little change in serum calcium, an increase in serum phosphate and a decrease in serum immunoreactive PTH concentration. They were classified as having suppressible hyperparathyroidism secondary to a primary renal calcium leak. Type 2b patients responded to both thiazide and 25(OH)D 3 therapy with an increase in serum calcium concentration and no decrease in immunoreactive PTH. They were classified as having nonsuppressible normocalcemic hyperparathyroidism and underwent successful parathyroid surgery. However, they had persistent postoperative hypercalciuria, and several had suppressible hyperparathyroidism several years after their successful parathyroid surgery. On the basis of these facts, type 2b patients were also considered to have a long-standing primary renal calcium leak that led initially to secondary hyperparathyroidism but eventually to tertiary or nonsuppressible hyperparathyroidism.

Aldosterone secretion: Effect of phorbol ester and A23187

The effects of the divalent ionophore, A23187, the phorbol ester, and/or 12-0-tetradecanoyl-phorbol-13-acetate on aldosterone secretion from adrenal glomerulosa cells were compared to those of angiotensin II (AII). AII causes a prompt and sustained increase in secretion. A23187 causes an initial increase followed by a gradual decline to values less than 25 percent of those seen with AII. TPA causes no initial increase but a slowly progressive rise in secretion rate to a less than maximal value. When TPA and A23187 act together, there is a prompt and sustained increase in aldosterone production rate similar to that seen after AII addition. The effect of TPA is dependent on the free Ca2+ concentration of the cell cytosol. These results are interpreted in terms of a model of cell activation in which two branches of the calcium messenger system operate to control respectively the initial and sustained phases of the secretory response. The first phase occurs as a consequence of amplitude modulation of the calmodulin branch of the system by a rise in [Ca2+]c, and the second phase as a consequence of the sensitivity modulation of the C-kinase branch by diacylglycerol.

Ionic and hormonal control of calcium homeostasis

Abstract Evidence obtained from analysis at a number of different levels of organization has led to the rather startling conclusion that parathyroid hormone (PTH) promotes bone resorption and inhibits bone formation by stimulating calcium uptake into cells and that calcitonin acts by stimulating calcium efflux from bone cells. In addition, the basis of PTH action on other cells appears to involve a similar type of control system. These hormonally induced changes in cellular calcium appear illogical at first glance and opposite to what one might have originally supposed. However, viewed from the perspective of evolution, and accepting the postulate that extracellular calcium homeostasis was achieved by adaptations of those ancient systems developed to maintain intracellular calcium homeostasis, they make considerable sense. In addition, insights gained from the study of the cellular basis of extracellular calcium homeostasis have led to the concept that the ancient system developed to achieve intracellular calcium homeostasis is a critically important part of the response of a wide variety of cells to specific stimuli. The unique features of extracellular calcium homeostasis thus lie not in unique cellular control systems but in the ability of specific hormones to activate selectively this ubiquitous cellular mechanism in certain highly differentiated cells in three distinct organs—gut, kidney and bone.

Physiology and Pathophysiology of Insulin Secretion

Mechanisms by which various classes of extracellular signals regulate insulin secretion are discussed regarding their cellular and molecular actions. Under physiological circumstances, the small postprandial changes in plasma glucose concentrations (∼4.4–6.6 mM) primarily serve as a conditional modifier of insulin secretion and dramatically alter the responsiveness of islets to a combination of neurohumoral agonists. These agonists have two functions. Cholecystokinin (CCK) and acetylcholine activate the hydrolysis of polyphosphoinositides, and gastric inhibitory polypeptide (GIP) and glucagonlike peptide 1 activate adenylate cyclase. These two functional classes of neurohumoral agonists act synergistically to enhance insulin secretion when plasma glucose is >6.0 mM but not when it is ≤4 mM. On the other hand, an increase in plasma glucose concentration to 8–10 mM induces an increase in insulin secretory rate in the absence of any of the neurohumoral agonists. Remarkably, high glucose leads to an increase in the same intracellular signals, as does a combination of acetylcholine and GIP. On the basis of these data, a model of how insulin secretion is regulated under physiological circumstances is proposed. This model emphasizes that the regulation of insulin secretion occurs in three stages: cephalic, early enteric, and later enteric. In this view, the crucial event occurring during the first two phases is the agonistinduced translocation of protein kinase C (PKC) to the plasma membrane under conditions in which an increase in Ca2+ influx does not occur. PKC is now in a cellular location and a Ca2+-sensitive conformation such that an increase in Ca2+ influx rate occurring during the third phase leads to its immediate activation and an enhanced rate of insulin secretion. Furthermore, under physiological circumstances, an optimal insulin secretory response is dependent on a correct temporal pattern of signals arising from neural and enteric sources. If this pattern is deranged, an abnormal pattern of insulin secretion is observed. An important new insight is provided by the observation that agonists (e.g., CCK or acetylcholine) that act to stimulate the hydrolysis of phosphatidylinositides, when acting for a short period (10–20 min), induce an enhanced responsiveness of islets to glucose, i.e., proemial sensitization. However, when acting unopposed for several hours, these agonists will induce a time-dependent suppression of responsiveness to glucose and other agonists. The latter observation implies that optimal insulin secretion is dependent on periodic rather than continuous exposure to the correct pattern of extracellular signals. The clinical implications of these new observations are discussed regarding glucose toxicity, the possible role of interleukin 1 in the pathogensis of insulin-dependent diabetes, sulfonylurea therapy, and the abnormalities of insulin secretion seen in non-insulin-dependent diabetes.