A monoclonal antibody against tyrosine hydroxylase: application in light and electron microscopy.

Semenenko, F. M.; Cuello, A. Claudio; Goldstein, Menek; Lee, K Y; Sidebottom, E

doi:10.1177/34.6.2871071

Cited by 31 publications

(13 citation statements)

References 11 publications

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“…In order to detect the TH immunoreactive (C) cell bodies, their immediate processes in the arcuate nucleus and distal axons in the median eminence free-floating sections were subjected to immunocytochemistry for TH as described by Ezquer & Seltzer (2003) using mouse monoclonal antibody against TH (Semenenko et al 1986) diluted in the ratio of 1:50 in carrier solution and revealed with avidin-biotin-peroxidase (ABC Kit, Vector Corporation) diluted to 1:100 in Tris buffer (TB, 0.1 M Tris-HCl (pH 7.4)). In the control specificity experiments, the primary antibody was omitted in the course of immunostaining.…”

Section: Th Ihcmentioning

confidence: 99%

“…The samples were run, transferred to nitrocellulose membranes (Hybond C Amershan Life Science), and blotted for TH immunoreactivity as described by Ezquer & Seltzer (2003) using the primary antibody anti-TH (Semenenko et al 1986), at 1:500 dilution and revealed with horseradish peroxidase-conjugated goat anti-mouse (DAKO Corporation, Carpinteria, CA, USA) and detection by chemiluminescence (ECL TM, Amersham Pharmacia Biotech). Multiple exposures of different times were made, to bring the exposures within the linear response range of the film (X-OMAT-AR, Kodak) and quantified by densitometry using digital image processing and the freeware program NIH Image 1.6/ppc (developed at the US National Institutes of Health and available on the internet at http://rsb.info.nih.gov/nih-image/).…”

Section: Th Western Blotsmentioning

confidence: 99%

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Hormonal profile and reproductive performance in lactation deficient (OFA hr/hr) and normal (Sprague–Dawley) female rats

Valdéz¹,

Penissi²,

Deis³

et al. 2007

Reproduction

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Lactation deficiency may have important consequences on infant health, particularly in populations of low socioeconomic status. The OFA hr/hr (OFA) strain of rats, derived from Sprague-Dawley (SD) rats, has deficient lactation and is a good model of lactation failure. We examined the reproductive performance and hormonal profiles in OFA and SD strains to determine the cause(s) of the lactation failure of the OFA strain. We measured hormonal (PRL, GH, gonadotropins, oxytocin, and progesterone) levels by RIA in cycling, pregnant, and lactating rats and in response to suckling. Dopaminergic metabolism was assessed by determination of mediobasal hypothalamic dopamine and dihydroxyphenylacetic acid (DOPAC) concentrations by HPLC and tyrosine hydroxylase expression by immunocytochemistry and western blot. OFA rats have normal fertility but 50% of the litters die of malnutrition on early lactation; only 6% of the mothers show normal lactation. The OFA rats showed lower circulating PRL during lactation, increased hypothalamic dopamine and DOPAC, and impaired milk ejection with decreased PRL and oxytocin response to suckling. Before parturition, PRL release and lactogenesis were normal, but dopaminergic metabolism was altered, suggesting activation of the dopaminergic system in OFA but not in SD rats. The number of arcuate and periventricular neurons expressing tyrosine hydroxylase was higher in SD rats, but hypothalamic expression of TH was higher in OFA rats at the end of pregnancy and early lactation. These results suggest that the OFA rats have impaired PRL release linked with an augmented dopaminergic tone which could be partially responsible for the lactational failure.

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Section: Th Ihcmentioning

confidence: 99%

Section: Th Western Blotsmentioning

confidence: 99%

Hormonal profile and reproductive performance in lactation deficient (OFA hr/hr) and normal (Sprague–Dawley) female rats

Valdéz¹,

Penissi²,

Deis³

et al. 2007

Reproduction

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“…The samples were analyzed for the presence of TH protein after separation by 12% SDS-PAGE on minigels in parallel with prestained protein molecular weight standards as described by Laemmli [35] loading 8 µg of cytoplasmatic proteins from each homogenate per lane. The samples were run, transferred to nitrocellulose membranes (Hybond C, Amershan Life Science) and blotted for TH immunoreactivity [29] using primary antibody anti-TH [34] at 1:500 dilution and revealed with horseradish peroxidase-conjugated goat anti-mouse (Dako Corp., Carpinteria, Calif., USA) and detection by chemiluminescence (ECL TM, Amersham Pharmacia Biotech, UK). Multiple exposures of different times were made, to bring the exposures within the linear response range of the film (X-OMAT-AR, Kodak) and quantified by densitometry using digital image processing and the freeware program NIH Image 1.6/ppc (developed at the US National Institutes of Health and available on the Internet at htpp://rsb.info.nih.gov/nih-image/).…”

Section: Methodsmentioning

confidence: 99%

“…In order to detect the TH-immunoreactive (+) cell bodies, their immediate processes in the arcuate nucleus and distal axons in the median eminence free-floating sections were subjected to immunocytochemistry for TH [29] using mouse monoclonal antibody against TH [34] diluted 1:50 in carrier solution and revealed with avidin biotin peroxidase diluted to 1:100 in Tris buffer (TB, 0.1 M Tris-HCl, pH 7.4) (ABC Kit, Vector Corp.). In the control specificity experiments the primary antibody was omitted in the course of immunostaining.…”

Section: Methodsmentioning

confidence: 99%

Dopaminergic Mechanisms Involved in Prolactin Release after Mifepristone and Naloxone Treatment during Late Pregnancy in the Rat

et al. 2006

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Background/Aims: During late pregnancy, the antiprogesterone mifepristone facilitates prolactin release. This effect is enhanced by administration of the opioid antagonist naloxone, suggesting an inhibitory-neuromodulatory role of the opioid system. Since hypothalamic dopamine (DA) is the main regulator of prolactin release, in this study we explored the role of DA on prolactin release induced by mifepristone and naloxone treatment. Methods/Results: Rats on day 19 of pregnancy were used. Naloxone treatment did not modify the 3,4-dihydroxyphenylacetic acid/DA (DOPAC/DA) ratio or serum prolactin concentration in control rats. After mifepristone treatment, DA activity diminished significantly without modifying serum prolactin levels. Naloxone administration to antiprogesterone-treated rats did not change the DOPAC/DA ratio but increased serum prolactin. Tyrosine hydroxylase (TH) expression in medial basal hypothalamus (MBH) protein extracts was lowered by pretreatment with mifepristone, with no additional effect of naloxone. While mifepristone decreased the intensity of TH immunoreactivity in the arcuate and periventricular nuclei and in fibers of the median eminence, naloxone treatment had no further effect. Conclusions: (1) A reduction of tuberoinfundibular dopaminergic (TIDA) neuron activity is suggested by the fall of the DOPAC/DA ratio and the low expression of MBH TH; (2) this reduction facilitates prolactin secretion by naloxone, indicating that progesterone stimulates DA neurons to maintain low serum prolactin; (3) naloxone action seems to depend on a previous decrease of DA tone induced by mifepristone, without involve a direct effect on neuronal DA activity, and (4) endogenous opioids may inhibit prolactin secretion through a non-dopaminergic neuronal system that regulates prolactin secretion in which as yet undetermined prolactin-releasing factors may participate.

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“…Therefore all catecholamine-producing neurones express tyrosine hydroxylase. Monoclonal antibodies against this enzyme have also been produced by Semenenko et al [6], Ross et al [7] and more recently by Rohrer et al [8], who used a monoclonal antibody reacting with tyrosine hydroxylase present in homogenates of quail, chicken, rat and bovine adrenal gland. Several laboratories have purified tyrosine hydroxylase and produced polyclonal antibodies, which have been used in immunohistochemical [2], immunotitration [3] and neuroenibryological [4] experiments as well as for the localization of tyrosine-hydroxylase-producing neurones in brain and chromaffin cells of adrenal glands [5].…”

mentioning

confidence: 99%

Role of the N‐terminus of rat pheochromocytoma tyrosine hydroxylase in the regulation of the enzyme's activity

Bonnefoy¹,

Ferrara²,

Rohrer³

et al. 1988

European Journal of Biochemistry

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Activation of rat pheochromocytoma tyrosine hydroxylase by limited tryptic proteolysis was investigated. The modifications produced upon the enzyme's structure were analyzed with the use of sodium dodecyl sulfate/polyacrylamide gel electrophoresis and tyrosine hydroxylase activity was measured all through the digestion. During the proteolysis the activity of tyrosine hydroxylase was elevated threefold at the same time as a 56-kDa tryptic fragment was formed. When the enzyme was phosphorylated, at its N-terminal region, by a kinase copurified with tyrosine hydroxylase, the major 56-kDa species did not appear to be phosphorylated on the autoradiograph, suggesting that it was derived from the native subunit by cleavage of the N-terminal of the protein. The reactivity of the 2/40/15 anti-(tyrosine hydroxylase) monoclonal antibody with the N-terminal of tyrosine hydroxylase was also investigated, using the Western-blot technique. This antibody reacted with the 62-kDa hydroxylase subunit but not with the 60-kDa tryptic fragment; the amino acid sequences of these two species showed that the 60-kDa fragment lacked the first 16 N-terminal amino acids of the native molecule. These results suggest that the N-terminal region of tyrosine hydroxylase is apparently responsible for an inhibition of the hydroxylase activity and that the first N-terminal amino acids of the hydroxylase are necessary for the recognition of the enzyme by its antibody.

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A monoclonal antibody against tyrosine hydroxylase: application in light and electron microscopy.

Cited by 31 publications

References 11 publications

Hormonal profile and reproductive performance in lactation deficient (OFA hr/hr) and normal (Sprague–Dawley) female rats

Hormonal profile and reproductive performance in lactation deficient (OFA hr/hr) and normal (Sprague–Dawley) female rats

Dopaminergic Mechanisms Involved in Prolactin Release after Mifepristone and Naloxone Treatment during Late Pregnancy in the Rat

Role of the N‐terminus of rat pheochromocytoma tyrosine hydroxylase in the regulation of the enzyme's activity

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