Journal of Toxicology and Environmental Health, 1984
A system of primary cultures of human chorionic trophoblasts has been used for studying the effec... more A system of primary cultures of human chorionic trophoblasts has been used for studying the effects of heavy metals on human reproductive tissue. Using this system, changes in cellular concentration of metallothionein (MT) in response to exposure to Cd or Zn were determined. Trophoblasts were isolated from term chorion leave, grown in RPMI-1640 medium, and exposed to Cd or Zn. Cellular content of MT was measured using the Cd/heme radioassay. MT increased in a concentration- and time-dependent manner after exposure to either metal. Cd increased the content of MT in trophoblasts at concentrations as low as 0.5 microM during a 24-h exposure. Moreover, extending exposure to Cd (2 microM) to 72 h resulted in a 3-4-fold increase in the concentration of MT. On a molar basis, Zn was not as potent a stimulus for MT synthesis as Cd, and required a concentration of 2.5 microM to increase the concentration of MT over a 24-h period. However, a 48- or 72-h exposure to Zn (10 microM) increased concentrations of MT nearly 8-fold over control values. Simultaneous exposure to Cd (2 microM) and inhibitors of protein synthesis, cycloheximide and actinomycin D, prevented the typical increase in MT concentration, suggesting that the metals act to increase the synthesis of MT. In another series of experiments, trophoblasts were exposed to Cd (2 microM) for 24 h, after which the cells were challenged with cytotoxic concentrations of Cd. Cells pretreated with Cd and then challenged with toxic concentrations of Cd had higher levels of MT and showed less toxicity, as indicated by leakage of lactic dehydrogenase. These results suggest that MT serves to sequester the metals in trophoblasts and reduce the toxicity of heavy metals. Thus, this system should be useful for studying the effects of heavy metals and characterizing the induction of MT in human reproductive tissue in vitro.
ABSTRACT Prorenin (Pro) is synthesized in a number of human utero-placental tissues, including ch... more ABSTRACT Prorenin (Pro) is synthesized in a number of human utero-placental tissues, including chorion, decidua, villous placenta and probably mesenchymal cells. The release of Pro from these extra-renal tissues follows new protein synthesis and appears to utilize the constitutive secretory pathway. Unlike processing in the kidney, very little of the Pro is subsequently cleaved to the smaller product (active renin). Primary signals which regulate Pro include protein hormones and peptides (relaxin, endothelin, hCG), amines (epinephrine, norepinephrine, and related beta adrenergic agents), and eicosanoids. These agents increase the mRNA for prorenin at a time before peak secretory effects are noted. Other extracellular signals have negative regulatory effects. These include angiotensin, endotoxin and cytokines (TNF-alpha and interleukin-1 B). There is also evidence that glucocorticoid receptor activation has an inhibitory effects on Pro release in placenta. Second messengers involved in the regulation of Pro include cyclic AMP and protein kinase A (PKA), protein kinase C (PKC), and calcium. The possible biological effect(s) of the extracellular Pro are unknown but may be due to direct generation of angiotensin I. Since angiotensin-peptides have a number of trophic effect on both vascular and non-vascular tissues, regulation of utero-placental Pro by autocrine, paracrine or endocrine signalling may be critical in normal fetal and/or placental development.
ABSTRACT Lung inflammation, vasculitis, and fibrosis occur in FES following bone fractures, surgi... more ABSTRACT Lung inflammation, vasculitis, and fibrosis occur in FES following bone fractures, surgical procedures, and liposuction. We reported the onset of myocardial damage in a rat model of FES with pulmonary fibrosis. This study continues our earlier work with attention to the evolution of the inflammatory process of the coronaries. Of 28 male SD rats, 14 received 0.2 mL IV triolein (FES model) and 14 received IV saline. Six weeks later, Losartan (0.3 mg/kg) was given IP to half of each group up to ten weeks. At necropsy, the tissues were stained for H&E, SMA-1, and Trichrome for collagen. Six photos at 400x were made of each heart. The coronary vasculitis was assessed by measurement of the medial thickening and lumen patency. In addition to the lung fibrosis, the heart of the triolein treated rats showed intense SMA-1 staining of the coronary media, intima, and adventitia. Lumen patency was reduced versus saline controls (p < 0.05). Losartan and saline treated rats did not differ from controls. Losartan protected the triolein treated rats from the heart damage with mild medial staining and wider lumen patency (p = 0.05). A small rim of collagen (trichrome) was seen in the adventitia of all the rats with more intensity in the triolein treated groups. Losartan reduced its presence. SMA-1 staining by evidencing the transformation of fibroblasts and myofibroblasts shows the beginnings of collagen-induced fibrosis. Would a longer treatment result in a more severe coronaropathy? The protective effect of Losartan was again confirmed underlining the role of the Renin-Angiotensin system in this disease. Information from this model may help treating patients with FES.
Purified fractions of chromaflin granules from the adrenal medullae of cats have been prepared fo... more Purified fractions of chromaflin granules from the adrenal medullae of cats have been prepared for an electron microscopic study of changes associated with the release of catecholamines. Fractions from unstimulated medullae consisted of electronopaque, membrane-bounded spheroidal granules of about 4500 A maximum diameter, similar to those seen in situ. Fractions from animals whose adrenal medullae had been stimulated with acetylcholine and depleted of catecholamines consisted almost entirely of electron-translucent structures. It is concluded that during release of catecholamines from the chrorrmffin granules, the granules discharge their electron-opaque contents, but not their membranes, which remain in the cell as discrete structures. RECENT chemical evidence suggests that stimulation of the chromaflin cells of the adrenal medulla causes catecholamines to be released directly from the well known chromaffin granules. Thus, in addition to catecholamines, two other constituents of chromaffin granules have been found in the effluents from secreting #ands in proportions similar to those found in the granules. These constituents are adenine nucleotides z-3 and proteins 4, 5 immunologically characteristic of the granules. Since phospholipid and cholesterol, lipids present in abundance in the membranes of chromaflin granules, do not escape into the effluents of perfused adrenal glands during catecholamine release, 6, 7 and since there is no fall in the lipids in subcellular fractions from catecholamine-depleted glands, 6, s it seems likely that after the granules discharge their contents their membranes are retained by the cells. Moreover, since density gradient centrifugation reveals that upon catecholamine depletion these lipids are found in a lighter layer, it has been suggested that the lipids are retained as discrete structures with reduced specific gravity, and further that these structures are probably the electron-translucent granules prominent in electron micrographs of stimulated chromaffln cells. 6, s The purpose of the present study was to examine this possibility.
Mechanisms of local regulation of blood flow through the extensive fetoplacental vascular bed of ... more Mechanisms of local regulation of blood flow through the extensive fetoplacental vascular bed of the mature human placenta are not understood. As the placenta is not innervated, there is no nervous control of vascular resistance. Locally-produced vasoactive autacoids such as prostaglandins and thromboxane could play a role in modulating fetoplacental blood flow, or humoral agents could be involved. Human placenta in the form of in vitro slice preparations has been shown to produce thromboxane B2, the stable metabolite of thromboxane A2, and to convert exogenous arachidonic acid to several prostanoids, including PGE2, PGD2, PGF2α and 6-keto-PGF1α a biologically inactive metabolite of prostacyclin, PGI2 (Mitchell et al., 1982; Makila et al., 1984; Harper et al., 1983). Also, higher levels of iPGE were found in umbilical venous plasma than in plasma from umbilical arteries (Bibby et al., 1979), suggesting that production of PGE by the placenta occurs in vivo. Thromboxane A2, PGE2, and PGI2 are known to have pressor or depressor actions in many vascular beds (Whittle and Moncada, 1984; Kadowitz et al., 1984). We therefore evaluated the effects of PGE2, PGF2α, PGI2, PGE1 and the endoperoxide analogue U46619, a thromboxane mimetic (Granstrom et al., 1983), on the resistance of human fetoplacental vasculature in isolated dual-perfused cotyledons from human term placenta. Furthermore, the formation of a potentially important humoral agent, angiotensin II (AII), within the fetoplacental vascular bed was studied.
Publisher Summary This chapter elaborates a study analyzing mechanisms of exocytosis. Exocytosis ... more Publisher Summary This chapter elaborates a study analyzing mechanisms of exocytosis. Exocytosis is a method of discharge of secretory products in which there is selective release of granule-bound material. Other forms of secretion may cause nonselective release of cellular constituents. In the adrenal medulla, for instance, elevated pH causes the release not only of catecholamines but also of a cytoplasmic marker, lactic dehydrogenase (LDH). This is probably indicative of cell damage. Reserpine can release catecholamines without discharge of other soluble granule constituents. This represents another mechanism of catecholamine release and is also different from exocytosis. Evidence that exocytosis occurs in the adrenal medulla in response to the physiological transmitter, acetylcholine, and other drugs have been reviewed. This chapter discusses some of the factors believed to be involved in exocytisis in the adrenal medulla. The chapter highlights that inhibitors of ATP synthesis increase the spontaneous release of catecholamines from the adrenal gland, while inhibiting the evoked release. These findings have been interpreted along with the evidence that ATP activates calcium uptake by cell organelles and that ATP can release catecholamines from isolated chromaffin granules.
The properties of renin granules isolated from rat renal cortex were studied. Renin granules were... more The properties of renin granules isolated from rat renal cortex were studied. Renin granules were thermolabile since in 10 min at 0°C twice as much renin was released as at +37°C. Addition of Ca++ (10‐6 M‐10‐2 M) did not affect the spontaneous release at +37°C, pH 6.5, during 10 or 30 min incubation. However, when pH was elevated to above 7, renin release was significantly increased by Ca++ (10‐3 M). Additions of various amounts of KCl, NaCl or MgCl2, which increased the osmolality less than 20 mOsm/kg, did not affect the stability of the renin granules. Mg‐ATP (0.5 and 5 mM) as well as Mg‐GTP (5 mM) stabilized renin granules at +37°C, pH 6.5, but the corresponding nitrogen analogues Mg‐AMP‐PNP and Mg‐GMP‐PNP (0.5 and 5 mM) were not effective. Neither did Mg‐AMP (5 mM) nor ATP (5 mM) without Mg++ affect the renin release. No stabilization was observed by Mg‐ATP and Mg‐GTP in the purified granule preparations. The results suggest the importance of the cleavage of the terminal phosphate in the stabilization process. When the granules prepared at 300 mOsm/kg were first kept at hyperosmotic medium (range 300–1650 mOsm/kg) and then moved back to 300 mOsm/kg, the granules tend to lyse the more the greater was the reduction of the osmolality. The granules were more stable in isotonic sucrose than in isotonic ionic medium.
Neurosecretory granules were isolated from bovine posterior pituitary glands and incubated at 30 ... more Neurosecretory granules were isolated from bovine posterior pituitary glands and incubated at 30 or 37° in a medium containing KCl and MgCl2. On exposure to ATP the granules released vasopressin, oxytocin, and protein. Vasopressin release in response to ATP was potentiated by phosphoenolpyruvate + pyruvate kinase, was inhibited by AMP, and was accompanied by a fall in the optical density (at 540 mµ) of the granule suspension. The neurosecretory granules possessed ATPase activity and ATP. It is suggested that ATP and ATPase may participate in the release of hormones from the intact posterior pituitary gland.
Neurosecretory granules prepared from bovine posterior pituitary glands by cell fractionation met... more Neurosecretory granules prepared from bovine posterior pituitary glands by cell fractionation methods contain adenosine triphosphate and adenosine triphosphatase activity. Addition of adenosine triphosphate to suspensions of granules stimulates release of vasopressin. It is suggested that adenosine triphosphate and adenosine triphosphatase participate in the storage and release of vasopressin.
... RELEASE FROM THE ADRENAL MEDULLA: EFFECT OF COLCHICINE, VINCA ALKALOIDS AND DEUTERIUM OXIDE&a... more ... RELEASE FROM THE ADRENAL MEDULLA: EFFECT OF COLCHICINE, VINCA ALKALOIDS AND DEUTERIUM OXIDE' ALAN M. POISNER AND JOSEPH BERNSTEIN' Department of Pharmacology, University of Kansas Medical Center, Kansas City, Kansas ...
NATURE 1299 branc, as suggested by Allison, which play their part in the extremely short lifespan... more NATURE 1299 branc, as suggested by Allison, which play their part in the extremely short lifespans of chick embryo erythrocytes. One of these factors might be the high degree of immaturity at which the chick embryo erythrocytes are poured out into the blood stream 9 • A full account of our calculations is to be published elsewhere 2 • Future investigations will show if this relatively small number of recirculations holds good also for mammalian primitive erythrocytes. There are indications that their average lifespans are also about one-quarter of that found in adult mammals 10 •
It is known that renin is present in fetal membranes, with the highest concentration in the chori... more It is known that renin is present in fetal membranes, with the highest concentration in the chorion laeve (reflected chorion). The purpose of this study was to identify and localize renin in human chorion laeve. Indirect immunofluorescent analysis, using antiserum against pure human kidney renin, revealed a single layer of cells in the chorion with strongly positive fluorescence. The presence of atrophic villi in this layer together with other morphological evidence indicate that the cells which are positive for renin are cytotrophoblasts. Isolated cells were prepared from the chorion by collagenase digestion, followed by filtration and density gradient centrifugation on Percoll. The isolated cells also showed a positive reaction with the immunofluorescent technique. Control experiments with nonimmune serum did not show fluorescent cells. Biochemical analysis using RIA of angiotensin I generated from sheep substrate indicated that most of the renin activity in the isolated cells was present as inactive renin (activated by trypsin). The presence of renin in trophoblastic cells may be of significance in local cardiovascular regulation, events associated with parturition, or pathophysiological manifestations of trophoblastic disease.
1. Cats' adrenal glands were acutely denervated and perfused, in situ, with Locke's solution. 2. ... more 1. Cats' adrenal glands were acutely denervated and perfused, in situ, with Locke's solution. 2. When the medullary secretogogue Ba (2-10 mm) was added to the perfusion medium, large amounts of AMP and traces of ATP were found in the venous effluent from the adrenal gland along with the catecholamines. In this respect Ba mimicked the physiological secretogogue, acetylcholine. 3. Control perfusions with known concentrations of ATP showed that under such conditions ATP was rapidly broken down within the adrenal vasculature. 4. When such intravascular hydrolysis of ATP was suppressed by perfusing with a Ca-free, Mg-free Locke's solution containing 1-2 mm EDTA, catecholamine secretion induced by Ba was accompanied by the discharge of large amounts of ATP but relatively little AMP. 5. The ATP that accompanied catecholamine secretion in such circumstances is assumed to derive from the 'heavy' nucleotide-rich chromaffin granules and it is concluded that the mechanism for catecholamine secretion does not depend on hydrolysis of the ATP within these granules. 6. The release of ATP (unhydrolysed) supports the hypothesis that the chromaffin granules discharge their contents at the cell surface by the process of 'reverse pinocytosis'.
Journal of Toxicology and Environmental Health, 1984
A system of primary cultures of human chorionic trophoblasts has been used for studying the effec... more A system of primary cultures of human chorionic trophoblasts has been used for studying the effects of heavy metals on human reproductive tissue. Using this system, changes in cellular concentration of metallothionein (MT) in response to exposure to Cd or Zn were determined. Trophoblasts were isolated from term chorion leave, grown in RPMI-1640 medium, and exposed to Cd or Zn. Cellular content of MT was measured using the Cd/heme radioassay. MT increased in a concentration- and time-dependent manner after exposure to either metal. Cd increased the content of MT in trophoblasts at concentrations as low as 0.5 microM during a 24-h exposure. Moreover, extending exposure to Cd (2 microM) to 72 h resulted in a 3-4-fold increase in the concentration of MT. On a molar basis, Zn was not as potent a stimulus for MT synthesis as Cd, and required a concentration of 2.5 microM to increase the concentration of MT over a 24-h period. However, a 48- or 72-h exposure to Zn (10 microM) increased concentrations of MT nearly 8-fold over control values. Simultaneous exposure to Cd (2 microM) and inhibitors of protein synthesis, cycloheximide and actinomycin D, prevented the typical increase in MT concentration, suggesting that the metals act to increase the synthesis of MT. In another series of experiments, trophoblasts were exposed to Cd (2 microM) for 24 h, after which the cells were challenged with cytotoxic concentrations of Cd. Cells pretreated with Cd and then challenged with toxic concentrations of Cd had higher levels of MT and showed less toxicity, as indicated by leakage of lactic dehydrogenase. These results suggest that MT serves to sequester the metals in trophoblasts and reduce the toxicity of heavy metals. Thus, this system should be useful for studying the effects of heavy metals and characterizing the induction of MT in human reproductive tissue in vitro.
ABSTRACT Prorenin (Pro) is synthesized in a number of human utero-placental tissues, including ch... more ABSTRACT Prorenin (Pro) is synthesized in a number of human utero-placental tissues, including chorion, decidua, villous placenta and probably mesenchymal cells. The release of Pro from these extra-renal tissues follows new protein synthesis and appears to utilize the constitutive secretory pathway. Unlike processing in the kidney, very little of the Pro is subsequently cleaved to the smaller product (active renin). Primary signals which regulate Pro include protein hormones and peptides (relaxin, endothelin, hCG), amines (epinephrine, norepinephrine, and related beta adrenergic agents), and eicosanoids. These agents increase the mRNA for prorenin at a time before peak secretory effects are noted. Other extracellular signals have negative regulatory effects. These include angiotensin, endotoxin and cytokines (TNF-alpha and interleukin-1 B). There is also evidence that glucocorticoid receptor activation has an inhibitory effects on Pro release in placenta. Second messengers involved in the regulation of Pro include cyclic AMP and protein kinase A (PKA), protein kinase C (PKC), and calcium. The possible biological effect(s) of the extracellular Pro are unknown but may be due to direct generation of angiotensin I. Since angiotensin-peptides have a number of trophic effect on both vascular and non-vascular tissues, regulation of utero-placental Pro by autocrine, paracrine or endocrine signalling may be critical in normal fetal and/or placental development.
ABSTRACT Lung inflammation, vasculitis, and fibrosis occur in FES following bone fractures, surgi... more ABSTRACT Lung inflammation, vasculitis, and fibrosis occur in FES following bone fractures, surgical procedures, and liposuction. We reported the onset of myocardial damage in a rat model of FES with pulmonary fibrosis. This study continues our earlier work with attention to the evolution of the inflammatory process of the coronaries. Of 28 male SD rats, 14 received 0.2 mL IV triolein (FES model) and 14 received IV saline. Six weeks later, Losartan (0.3 mg/kg) was given IP to half of each group up to ten weeks. At necropsy, the tissues were stained for H&E, SMA-1, and Trichrome for collagen. Six photos at 400x were made of each heart. The coronary vasculitis was assessed by measurement of the medial thickening and lumen patency. In addition to the lung fibrosis, the heart of the triolein treated rats showed intense SMA-1 staining of the coronary media, intima, and adventitia. Lumen patency was reduced versus saline controls (p < 0.05). Losartan and saline treated rats did not differ from controls. Losartan protected the triolein treated rats from the heart damage with mild medial staining and wider lumen patency (p = 0.05). A small rim of collagen (trichrome) was seen in the adventitia of all the rats with more intensity in the triolein treated groups. Losartan reduced its presence. SMA-1 staining by evidencing the transformation of fibroblasts and myofibroblasts shows the beginnings of collagen-induced fibrosis. Would a longer treatment result in a more severe coronaropathy? The protective effect of Losartan was again confirmed underlining the role of the Renin-Angiotensin system in this disease. Information from this model may help treating patients with FES.
Purified fractions of chromaflin granules from the adrenal medullae of cats have been prepared fo... more Purified fractions of chromaflin granules from the adrenal medullae of cats have been prepared for an electron microscopic study of changes associated with the release of catecholamines. Fractions from unstimulated medullae consisted of electronopaque, membrane-bounded spheroidal granules of about 4500 A maximum diameter, similar to those seen in situ. Fractions from animals whose adrenal medullae had been stimulated with acetylcholine and depleted of catecholamines consisted almost entirely of electron-translucent structures. It is concluded that during release of catecholamines from the chrorrmffin granules, the granules discharge their electron-opaque contents, but not their membranes, which remain in the cell as discrete structures. RECENT chemical evidence suggests that stimulation of the chromaflin cells of the adrenal medulla causes catecholamines to be released directly from the well known chromaffin granules. Thus, in addition to catecholamines, two other constituents of chromaffin granules have been found in the effluents from secreting #ands in proportions similar to those found in the granules. These constituents are adenine nucleotides z-3 and proteins 4, 5 immunologically characteristic of the granules. Since phospholipid and cholesterol, lipids present in abundance in the membranes of chromaflin granules, do not escape into the effluents of perfused adrenal glands during catecholamine release, 6, 7 and since there is no fall in the lipids in subcellular fractions from catecholamine-depleted glands, 6, s it seems likely that after the granules discharge their contents their membranes are retained by the cells. Moreover, since density gradient centrifugation reveals that upon catecholamine depletion these lipids are found in a lighter layer, it has been suggested that the lipids are retained as discrete structures with reduced specific gravity, and further that these structures are probably the electron-translucent granules prominent in electron micrographs of stimulated chromaffln cells. 6, s The purpose of the present study was to examine this possibility.
Mechanisms of local regulation of blood flow through the extensive fetoplacental vascular bed of ... more Mechanisms of local regulation of blood flow through the extensive fetoplacental vascular bed of the mature human placenta are not understood. As the placenta is not innervated, there is no nervous control of vascular resistance. Locally-produced vasoactive autacoids such as prostaglandins and thromboxane could play a role in modulating fetoplacental blood flow, or humoral agents could be involved. Human placenta in the form of in vitro slice preparations has been shown to produce thromboxane B2, the stable metabolite of thromboxane A2, and to convert exogenous arachidonic acid to several prostanoids, including PGE2, PGD2, PGF2α and 6-keto-PGF1α a biologically inactive metabolite of prostacyclin, PGI2 (Mitchell et al., 1982; Makila et al., 1984; Harper et al., 1983). Also, higher levels of iPGE were found in umbilical venous plasma than in plasma from umbilical arteries (Bibby et al., 1979), suggesting that production of PGE by the placenta occurs in vivo. Thromboxane A2, PGE2, and PGI2 are known to have pressor or depressor actions in many vascular beds (Whittle and Moncada, 1984; Kadowitz et al., 1984). We therefore evaluated the effects of PGE2, PGF2α, PGI2, PGE1 and the endoperoxide analogue U46619, a thromboxane mimetic (Granstrom et al., 1983), on the resistance of human fetoplacental vasculature in isolated dual-perfused cotyledons from human term placenta. Furthermore, the formation of a potentially important humoral agent, angiotensin II (AII), within the fetoplacental vascular bed was studied.
Publisher Summary This chapter elaborates a study analyzing mechanisms of exocytosis. Exocytosis ... more Publisher Summary This chapter elaborates a study analyzing mechanisms of exocytosis. Exocytosis is a method of discharge of secretory products in which there is selective release of granule-bound material. Other forms of secretion may cause nonselective release of cellular constituents. In the adrenal medulla, for instance, elevated pH causes the release not only of catecholamines but also of a cytoplasmic marker, lactic dehydrogenase (LDH). This is probably indicative of cell damage. Reserpine can release catecholamines without discharge of other soluble granule constituents. This represents another mechanism of catecholamine release and is also different from exocytosis. Evidence that exocytosis occurs in the adrenal medulla in response to the physiological transmitter, acetylcholine, and other drugs have been reviewed. This chapter discusses some of the factors believed to be involved in exocytisis in the adrenal medulla. The chapter highlights that inhibitors of ATP synthesis increase the spontaneous release of catecholamines from the adrenal gland, while inhibiting the evoked release. These findings have been interpreted along with the evidence that ATP activates calcium uptake by cell organelles and that ATP can release catecholamines from isolated chromaffin granules.
The properties of renin granules isolated from rat renal cortex were studied. Renin granules were... more The properties of renin granules isolated from rat renal cortex were studied. Renin granules were thermolabile since in 10 min at 0°C twice as much renin was released as at +37°C. Addition of Ca++ (10‐6 M‐10‐2 M) did not affect the spontaneous release at +37°C, pH 6.5, during 10 or 30 min incubation. However, when pH was elevated to above 7, renin release was significantly increased by Ca++ (10‐3 M). Additions of various amounts of KCl, NaCl or MgCl2, which increased the osmolality less than 20 mOsm/kg, did not affect the stability of the renin granules. Mg‐ATP (0.5 and 5 mM) as well as Mg‐GTP (5 mM) stabilized renin granules at +37°C, pH 6.5, but the corresponding nitrogen analogues Mg‐AMP‐PNP and Mg‐GMP‐PNP (0.5 and 5 mM) were not effective. Neither did Mg‐AMP (5 mM) nor ATP (5 mM) without Mg++ affect the renin release. No stabilization was observed by Mg‐ATP and Mg‐GTP in the purified granule preparations. The results suggest the importance of the cleavage of the terminal phosphate in the stabilization process. When the granules prepared at 300 mOsm/kg were first kept at hyperosmotic medium (range 300–1650 mOsm/kg) and then moved back to 300 mOsm/kg, the granules tend to lyse the more the greater was the reduction of the osmolality. The granules were more stable in isotonic sucrose than in isotonic ionic medium.
Neurosecretory granules were isolated from bovine posterior pituitary glands and incubated at 30 ... more Neurosecretory granules were isolated from bovine posterior pituitary glands and incubated at 30 or 37° in a medium containing KCl and MgCl2. On exposure to ATP the granules released vasopressin, oxytocin, and protein. Vasopressin release in response to ATP was potentiated by phosphoenolpyruvate + pyruvate kinase, was inhibited by AMP, and was accompanied by a fall in the optical density (at 540 mµ) of the granule suspension. The neurosecretory granules possessed ATPase activity and ATP. It is suggested that ATP and ATPase may participate in the release of hormones from the intact posterior pituitary gland.
Neurosecretory granules prepared from bovine posterior pituitary glands by cell fractionation met... more Neurosecretory granules prepared from bovine posterior pituitary glands by cell fractionation methods contain adenosine triphosphate and adenosine triphosphatase activity. Addition of adenosine triphosphate to suspensions of granules stimulates release of vasopressin. It is suggested that adenosine triphosphate and adenosine triphosphatase participate in the storage and release of vasopressin.
... RELEASE FROM THE ADRENAL MEDULLA: EFFECT OF COLCHICINE, VINCA ALKALOIDS AND DEUTERIUM OXIDE&a... more ... RELEASE FROM THE ADRENAL MEDULLA: EFFECT OF COLCHICINE, VINCA ALKALOIDS AND DEUTERIUM OXIDE' ALAN M. POISNER AND JOSEPH BERNSTEIN' Department of Pharmacology, University of Kansas Medical Center, Kansas City, Kansas ...
NATURE 1299 branc, as suggested by Allison, which play their part in the extremely short lifespan... more NATURE 1299 branc, as suggested by Allison, which play their part in the extremely short lifespans of chick embryo erythrocytes. One of these factors might be the high degree of immaturity at which the chick embryo erythrocytes are poured out into the blood stream 9 • A full account of our calculations is to be published elsewhere 2 • Future investigations will show if this relatively small number of recirculations holds good also for mammalian primitive erythrocytes. There are indications that their average lifespans are also about one-quarter of that found in adult mammals 10 •
It is known that renin is present in fetal membranes, with the highest concentration in the chori... more It is known that renin is present in fetal membranes, with the highest concentration in the chorion laeve (reflected chorion). The purpose of this study was to identify and localize renin in human chorion laeve. Indirect immunofluorescent analysis, using antiserum against pure human kidney renin, revealed a single layer of cells in the chorion with strongly positive fluorescence. The presence of atrophic villi in this layer together with other morphological evidence indicate that the cells which are positive for renin are cytotrophoblasts. Isolated cells were prepared from the chorion by collagenase digestion, followed by filtration and density gradient centrifugation on Percoll. The isolated cells also showed a positive reaction with the immunofluorescent technique. Control experiments with nonimmune serum did not show fluorescent cells. Biochemical analysis using RIA of angiotensin I generated from sheep substrate indicated that most of the renin activity in the isolated cells was present as inactive renin (activated by trypsin). The presence of renin in trophoblastic cells may be of significance in local cardiovascular regulation, events associated with parturition, or pathophysiological manifestations of trophoblastic disease.
1. Cats' adrenal glands were acutely denervated and perfused, in situ, with Locke's solution. 2. ... more 1. Cats' adrenal glands were acutely denervated and perfused, in situ, with Locke's solution. 2. When the medullary secretogogue Ba (2-10 mm) was added to the perfusion medium, large amounts of AMP and traces of ATP were found in the venous effluent from the adrenal gland along with the catecholamines. In this respect Ba mimicked the physiological secretogogue, acetylcholine. 3. Control perfusions with known concentrations of ATP showed that under such conditions ATP was rapidly broken down within the adrenal vasculature. 4. When such intravascular hydrolysis of ATP was suppressed by perfusing with a Ca-free, Mg-free Locke's solution containing 1-2 mm EDTA, catecholamine secretion induced by Ba was accompanied by the discharge of large amounts of ATP but relatively little AMP. 5. The ATP that accompanied catecholamine secretion in such circumstances is assumed to derive from the 'heavy' nucleotide-rich chromaffin granules and it is concluded that the mechanism for catecholamine secretion does not depend on hydrolysis of the ATP within these granules. 6. The release of ATP (unhydrolysed) supports the hypothesis that the chromaffin granules discharge their contents at the cell surface by the process of 'reverse pinocytosis'.
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