Archives of Environmental Health: An International Journal
ISSN: 0003-9896 (Print) (Online) Journal homepage: http://www.tandfonline.com/loi/vzeh20
Ammonium and Sulfate Ion Release of Histamine From Lung Fragments Jeffrey M. Charles & Daniel B. Menzel PhD To cite this article: Jeffrey M. Charles & Daniel B. Menzel PhD (1975) Ammonium and Sulfate Ion Release of Histamine From Lung Fragments, Archives of Environmental Health: An International Journal, 30:6, 314-316, DOI: 10.1080/00039896.1975.10666706 To link to this article: http://dx.doi.org/10.1080/00039896.1975.10666706
Published online: 02 May 2013.
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Date: 28 June 2016, At: 16:32
Ammonium and Sulfate Ion Release of Histamine From Lung Fragments Downloaded by [University of California, San Diego] at 16:32 28 June 2016
Jeffrey M. Charles, Daniel B. Menzel, PhD
In vitro studies with gUinea pig lung fragments incubated with 10- to 200-mM concentrations of ammonium ion demonstrated the release of substantial quantities of histamine. Of the anions tested with ammonium ion, sulfate was the most potent, while nitrate and acetate ions were of intermediate potency and chloride was less potent. An osmotic effect is unlikely since equal concentrations of sodium chloride failed to release histamine. Isoproterenol, known to decrease anaphylactic histamine release, and acetylcholine, known to increase histamine release, had no effect on the ammonium sulfate-mediated release of histamine. N",2'-0-Dibutyryl adenosine 3',5' monophosphate (dibutyryl c-AMP) was also ineffective. These studies suggest that the inhalation irritation associated with certain sulfate and other salts, may be a function of their ability to release histamine in the presence of ammonium ion.
S
tudies of the chemistry of the atmosphere have shown that a portion of the sulfur dioxide emitted into the atmosphere undergoes oxidation, which leads to the formation of sulfuric acid and sulfate particulates. Amdur,l has provided data demonstrating that some of these particulate sulfur oxidation products have a greater irritant potential than sulfur dioxide itself. McJilton et aP showed that sulfur dioxide became more irritating as humidity increased, with the suggestion that sulSubmitted for publication Sept 23, 1974; accepted Oct 21. From the departments of physiology and pharmacology, and medicine, Duke Medical Center, Durham, NC. Reprint requests to the Division of Pharmacology, Duke Medical Center, PO Box 3709, Durham NC 27710 (Dr. Menzel).
314 Arch Environ Health/Vol 30, June 1975
furic acid or sulfate was responsible for the irritation. Catalytic converters, to be installed on all new automobiles in 1975, may become a new source of increased sulfates and sulfuric acid mist in urban areas. The observations of Amdur and Corn,~ that ammonium sulfate was highly irritating, led us to examine the effects of ammonium sulfate and other salts on the histamine release directly from fragments of guinea pig lung in vitro. METHODS AND MATERIALS Male Camm-Hartley guinea pigs weighing between 300 to 400 gm, were anesthetized and the lungs excised. The lungs were cut into fragments (75 to 150 mg), which were washed repeatedly with Tyrode's solution until the fragments were free of blood. The Tyrode's solution con~ tained 0.9 gm NaCI, 0.02 gm KCI, 0.02 gm CaCl z, 0.01 gm MgCl 2 , 0.1 gm glucose, 0.1 gm NaHCO" and 0.005 gm NaH 2 PO,1l00 ml. One or two lung fragments were employed in each test as a sample weighing 150 to 200 mg. The samples were then suspended in 3.0 ml of Tyrode's solution (pH 7.4) that contained varied concentrations (10 to 200 mp) of the various salts under study. The pH of all the solutions was carefully controlled and there was no modification in the pH as a result of the addition of the salts studied. The fragments were incubated in a shaking water bath at 37 C for 30 minutes, and the supernatant was assayed for the histamine released. The salts studied in this manner were sodium chloride, sodium sulfate, ammonium chloride, ammonium sulfate, ammonium nitrate, and ammonium citrate, Isoproterenol and acetylcholine were tested by preincubating the fragments for five minutes with 10- 3 and 10-1-M concentrations, re-
spectively, before the addition of the salts. Dibutyryl c-AMP was tested in a similar manner at 10"" M. Total histamine (ttg/gm of lung) was determined by boiling fresh lung slices for eight minutes and assaying the supernatant. Histamine Assay
Histamine was measured spectrophotofluorometrically,'1.5 A total of 0.2 ml of 70% perchloric acid was added to 2.0 ml of sample solution and the mixture incubated for 30 minutes. One milliliter of the supernatant was added to 0.75 gm NaCI, and then 1.5 ml n-butanol and 0.3 ml5N NaOH were added and mixed. The lower aqueous phase was removed by suction and 1.5 ml IN NaOH saturated with NaCl was added to the remaining phase and mixed. One milliliter of the upper butanol layer was transferred to 1.5 ml hexane and 1.25 ml O.lN HCI and the mixture agitated. The upper organic phase was removed by suction and 0.45 ml of the acid phase was added to 0.1 NaOH and 0.05 ml of a 0.5% methanolic solution of o-phthaldialdehyde. The reaction was stopped after four minutes by the addition of 0.05 ml 3N HCI. After the addition of 1.5 ml distilled water, the fluorescence was measured at 450 nm by excitation at 360 nm.
RESULTS
Lung fragments incubated with ammonium sulfate, ammonium nitrate, ammonium acetate, and ammonium chloride, in concentrations of 10 to 200 mM, released histamine in proportion to the concentration of the salts present (Figure). Sodium sulfate and sodium chloride, however, did not release any detectable histamine. Those salts that did release histamine varied in efficacy. The most efficacious, ammonium sulfate, produced Histamine Release/Charles & Menzel
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30 Table i.-Release of Histamine From Lung Fragments in the Presence of i00-mM Salt Solutions
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20 Compound Sodium chloride Sodium sulfate Ammonium chloride Ammonium acetate Ammonium nitrate sulfate "
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ISSN: 0003-9896 (Print) (Online) Journal homepage: http://www.tandfonline.com/loi/vzeh20
Ammonium and Sulfate Ion Release of Histamine From Lung Fragments Jeffrey M. Charles & Daniel B. Menzel PhD To cite this article: Jeffrey M. Charles & Daniel B. Menzel PhD (1975) Ammonium and Sulfate Ion Release of Histamine From Lung Fragments, Archives of Environmental Health: An International Journal, 30:6, 314-316, DOI: 10.1080/00039896.1975.10666706 To link to this article: http://dx.doi.org/10.1080/00039896.1975.10666706
Published online: 02 May 2013.
Submit your article to this journal
Article views: 1
View related articles
Citing articles: 3 View citing articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=vzeh20 Download by: [University of California, San Diego]
Date: 28 June 2016, At: 16:32
Ammonium and Sulfate Ion Release of Histamine From Lung Fragments Downloaded by [University of California, San Diego] at 16:32 28 June 2016
Jeffrey M. Charles, Daniel B. Menzel, PhD
In vitro studies with gUinea pig lung fragments incubated with 10- to 200-mM concentrations of ammonium ion demonstrated the release of substantial quantities of histamine. Of the anions tested with ammonium ion, sulfate was the most potent, while nitrate and acetate ions were of intermediate potency and chloride was less potent. An osmotic effect is unlikely since equal concentrations of sodium chloride failed to release histamine. Isoproterenol, known to decrease anaphylactic histamine release, and acetylcholine, known to increase histamine release, had no effect on the ammonium sulfate-mediated release of histamine. N",2'-0-Dibutyryl adenosine 3',5' monophosphate (dibutyryl c-AMP) was also ineffective. These studies suggest that the inhalation irritation associated with certain sulfate and other salts, may be a function of their ability to release histamine in the presence of ammonium ion.
S
tudies of the chemistry of the atmosphere have shown that a portion of the sulfur dioxide emitted into the atmosphere undergoes oxidation, which leads to the formation of sulfuric acid and sulfate particulates. Amdur,l has provided data demonstrating that some of these particulate sulfur oxidation products have a greater irritant potential than sulfur dioxide itself. McJilton et aP showed that sulfur dioxide became more irritating as humidity increased, with the suggestion that sulSubmitted for publication Sept 23, 1974; accepted Oct 21. From the departments of physiology and pharmacology, and medicine, Duke Medical Center, Durham, NC. Reprint requests to the Division of Pharmacology, Duke Medical Center, PO Box 3709, Durham NC 27710 (Dr. Menzel).
314 Arch Environ Health/Vol 30, June 1975
furic acid or sulfate was responsible for the irritation. Catalytic converters, to be installed on all new automobiles in 1975, may become a new source of increased sulfates and sulfuric acid mist in urban areas. The observations of Amdur and Corn,~ that ammonium sulfate was highly irritating, led us to examine the effects of ammonium sulfate and other salts on the histamine release directly from fragments of guinea pig lung in vitro. METHODS AND MATERIALS Male Camm-Hartley guinea pigs weighing between 300 to 400 gm, were anesthetized and the lungs excised. The lungs were cut into fragments (75 to 150 mg), which were washed repeatedly with Tyrode's solution until the fragments were free of blood. The Tyrode's solution con~ tained 0.9 gm NaCI, 0.02 gm KCI, 0.02 gm CaCl z, 0.01 gm MgCl 2 , 0.1 gm glucose, 0.1 gm NaHCO" and 0.005 gm NaH 2 PO,1l00 ml. One or two lung fragments were employed in each test as a sample weighing 150 to 200 mg. The samples were then suspended in 3.0 ml of Tyrode's solution (pH 7.4) that contained varied concentrations (10 to 200 mp) of the various salts under study. The pH of all the solutions was carefully controlled and there was no modification in the pH as a result of the addition of the salts studied. The fragments were incubated in a shaking water bath at 37 C for 30 minutes, and the supernatant was assayed for the histamine released. The salts studied in this manner were sodium chloride, sodium sulfate, ammonium chloride, ammonium sulfate, ammonium nitrate, and ammonium citrate, Isoproterenol and acetylcholine were tested by preincubating the fragments for five minutes with 10- 3 and 10-1-M concentrations, re-
spectively, before the addition of the salts. Dibutyryl c-AMP was tested in a similar manner at 10"" M. Total histamine (ttg/gm of lung) was determined by boiling fresh lung slices for eight minutes and assaying the supernatant. Histamine Assay
Histamine was measured spectrophotofluorometrically,'1.5 A total of 0.2 ml of 70% perchloric acid was added to 2.0 ml of sample solution and the mixture incubated for 30 minutes. One milliliter of the supernatant was added to 0.75 gm NaCI, and then 1.5 ml n-butanol and 0.3 ml5N NaOH were added and mixed. The lower aqueous phase was removed by suction and 1.5 ml IN NaOH saturated with NaCl was added to the remaining phase and mixed. One milliliter of the upper butanol layer was transferred to 1.5 ml hexane and 1.25 ml O.lN HCI and the mixture agitated. The upper organic phase was removed by suction and 0.45 ml of the acid phase was added to 0.1 NaOH and 0.05 ml of a 0.5% methanolic solution of o-phthaldialdehyde. The reaction was stopped after four minutes by the addition of 0.05 ml 3N HCI. After the addition of 1.5 ml distilled water, the fluorescence was measured at 450 nm by excitation at 360 nm.
RESULTS
Lung fragments incubated with ammonium sulfate, ammonium nitrate, ammonium acetate, and ammonium chloride, in concentrations of 10 to 200 mM, released histamine in proportion to the concentration of the salts present (Figure). Sodium sulfate and sodium chloride, however, did not release any detectable histamine. Those salts that did release histamine varied in efficacy. The most efficacious, ammonium sulfate, produced Histamine Release/Charles & Menzel
'0 z =>
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30 Table i.-Release of Histamine From Lung Fragments in the Presence of i00-mM Salt Solutions
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20 Compound Sodium chloride Sodium sulfate Ammonium chloride Ammonium acetate Ammonium nitrate sulfate "
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