Scintillation autoradiographic localization of 1,25-dihydroxyvitamin D3 in chick intestine

P. G. Jones, Mark R Haussler

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

The intracellular binding site of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] was determined via preliminary biochemical analysis of radioactive 1,25(OH)2D3 association with various chick tissues and then by direct autoradiographic visualization. When vitamin D-deficient chicks were injected intracardially with physiological doses of tritiated 1,25(OH)2D3 and killed 2 h later, 2-3 times more radioactivity was found in the intestinal mucosa than was present in equal weights of pancreas, parathyroid, or liver tissue. Very little tritium was found in muscle tissue. The intestinally localized radioactivity was predominantly associated with the nuclear chromatin fraction, and binding of 1,25(OH)2[3H]D3 to the nucleus was maximal 2 h after injection and at a dose of a least 0.52 nmol. Using this dose and time period, autoradiographic studies were done on duodenum and thoracic muscle of rachitic chicks injected with radioactive 1,25(OH)2D3 (11.2 Ci/mol). Ci/mol. Thin sections of tissue were prepared for thaw and dry mount scintillation autoradiography as well as simple dip-coating autoradiography. After exposure for 4-6 months, a preferential concentration and retention of tritiumlabeled 1,25(OH)2D3 was evident in the nuclei of intestinal villi and in the crypt of Lieberkuehn cells when each of the autoradiographic techniques was utilized. Quantitation of the labeled hormone by counting the number of nuclear and cytoplasmic silver grains in target intestinal tissue per 500 μm2 area confirms the significant nuclear accumulation in both villi and crypt cells. No such nuclear concentration of silver grains was observed in thoracic muscle cells, and the intestinal localization was abolished when a 100-fold excess of unlabeled 1,25(OH)2D3 was injected simultaneously with the radioactive hormone. Thus, from these direct observations in morphologically intact tissue, we conclude that 1,25(OH)2D3 is bound in a tissue-selective fashion to a high affinity, low capacity site within the nucleus of its intestinal target organ.

Original languageEnglish (US)
Pages (from-to)313-321
Number of pages9
JournalEndocrinology
Volume104
Issue number2
StatePublished - 1979

Fingerprint

Calcitriol
Intestines
Autoradiography
Silver
Radioactivity
Thorax
Hormones
Muscles
Rickets
Tritium
Intestinal Mucosa
Duodenum
Vitamin D
Muscle Cells
Chromatin
Pancreas
Binding Sites
Weights and Measures
Injections
Liver

ASJC Scopus subject areas

  • Endocrinology
  • Endocrinology, Diabetes and Metabolism

Cite this

Scintillation autoradiographic localization of 1,25-dihydroxyvitamin D3 in chick intestine. / Jones, P. G.; Haussler, Mark R.

In: Endocrinology, Vol. 104, No. 2, 1979, p. 313-321.

Research output: Contribution to journalArticle

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abstract = "The intracellular binding site of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] was determined via preliminary biochemical analysis of radioactive 1,25(OH)2D3 association with various chick tissues and then by direct autoradiographic visualization. When vitamin D-deficient chicks were injected intracardially with physiological doses of tritiated 1,25(OH)2D3 and killed 2 h later, 2-3 times more radioactivity was found in the intestinal mucosa than was present in equal weights of pancreas, parathyroid, or liver tissue. Very little tritium was found in muscle tissue. The intestinally localized radioactivity was predominantly associated with the nuclear chromatin fraction, and binding of 1,25(OH)2[3H]D3 to the nucleus was maximal 2 h after injection and at a dose of a least 0.52 nmol. Using this dose and time period, autoradiographic studies were done on duodenum and thoracic muscle of rachitic chicks injected with radioactive 1,25(OH)2D3 (11.2 Ci/mol). Ci/mol. Thin sections of tissue were prepared for thaw and dry mount scintillation autoradiography as well as simple dip-coating autoradiography. After exposure for 4-6 months, a preferential concentration and retention of tritiumlabeled 1,25(OH)2D3 was evident in the nuclei of intestinal villi and in the crypt of Lieberkuehn cells when each of the autoradiographic techniques was utilized. Quantitation of the labeled hormone by counting the number of nuclear and cytoplasmic silver grains in target intestinal tissue per 500 μm2 area confirms the significant nuclear accumulation in both villi and crypt cells. No such nuclear concentration of silver grains was observed in thoracic muscle cells, and the intestinal localization was abolished when a 100-fold excess of unlabeled 1,25(OH)2D3 was injected simultaneously with the radioactive hormone. Thus, from these direct observations in morphologically intact tissue, we conclude that 1,25(OH)2D3 is bound in a tissue-selective fashion to a high affinity, low capacity site within the nucleus of its intestinal target organ.",
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AB - The intracellular binding site of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] was determined via preliminary biochemical analysis of radioactive 1,25(OH)2D3 association with various chick tissues and then by direct autoradiographic visualization. When vitamin D-deficient chicks were injected intracardially with physiological doses of tritiated 1,25(OH)2D3 and killed 2 h later, 2-3 times more radioactivity was found in the intestinal mucosa than was present in equal weights of pancreas, parathyroid, or liver tissue. Very little tritium was found in muscle tissue. The intestinally localized radioactivity was predominantly associated with the nuclear chromatin fraction, and binding of 1,25(OH)2[3H]D3 to the nucleus was maximal 2 h after injection and at a dose of a least 0.52 nmol. Using this dose and time period, autoradiographic studies were done on duodenum and thoracic muscle of rachitic chicks injected with radioactive 1,25(OH)2D3 (11.2 Ci/mol). Ci/mol. Thin sections of tissue were prepared for thaw and dry mount scintillation autoradiography as well as simple dip-coating autoradiography. After exposure for 4-6 months, a preferential concentration and retention of tritiumlabeled 1,25(OH)2D3 was evident in the nuclei of intestinal villi and in the crypt of Lieberkuehn cells when each of the autoradiographic techniques was utilized. Quantitation of the labeled hormone by counting the number of nuclear and cytoplasmic silver grains in target intestinal tissue per 500 μm2 area confirms the significant nuclear accumulation in both villi and crypt cells. No such nuclear concentration of silver grains was observed in thoracic muscle cells, and the intestinal localization was abolished when a 100-fold excess of unlabeled 1,25(OH)2D3 was injected simultaneously with the radioactive hormone. Thus, from these direct observations in morphologically intact tissue, we conclude that 1,25(OH)2D3 is bound in a tissue-selective fashion to a high affinity, low capacity site within the nucleus of its intestinal target organ.

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