Urocortin-like Immunoreactivity Research Articles

Increased oxytocin-monomeric red fluorescent protein 1 fluorescent intensity with urocortin-like immunoreactivity in the hypothalamo-neurohypophysial system of aged transgenic rats

To visualize oxytocin in the hypothalamo-neurohypophysial system, we generated a transgenic rat that expresses the oxytocin-monomeric red fluorescent protein 1 (mRFP1) fusion gene. In the present study, we examined the age-related changes of oxytocin-mRFP1 fluorescent intensity in the posterior pituitary (PP), the supraoptic nucleus (SON) and the paraventricular nucleus (PVN) of transgenic rats. The mRFP1 fluorescent intensities were significantly increased in the PP, the SON and the PVN of 12-, 18- and 24-month-old transgenic rats in comparison with 3-month-old transgenic rats. Immunohistochemical staining for urocortin, which belongs to the family of corticotropin-releasing factor family, revealed that the numbers of urocortin-like immunoreactive (LI) cells in the SON and the PVN were significantly increased in 12-, 18- and 24-month-old transgenic rats in comparison with 3-month-old transgenic rats. Almost all of urocortin-LI cells co-exist mRFP1-expressing cells in the SON and the PVN of aged transgenic rats. These results suggest that oxytocin content of the hypothalamo-neurohypophysial system may be modulated by age-related regulation. The physiological role of the co-existence of oxytocin and urocortin in the SON and PVN of aged rats remains unclear.

Urocortin-like immunoreactivity in the primary lymphoid organs of the duck (Anas platyrhynchos)

Urocortin (UCN) is a 40 aminoacid peptide which belongs to corticotropin-releasing factor (CRF) family. This family of peptides stimulates the secretion of proopiomelanocortin (POMC)-derived peptides, adrenocorticotropic hormone (ACTH), beta-endorphin and melanocyte-stimulating hormone (MSH) in the pituitary gland. In the present study, using Western blotting and immunohistochemistry, the distribution of UCN in the primary lymphoid organs of the duck was investigated at different ages. In the cloacal burse and thymus, Western blot demonstrated the presence of a peptide having a molecular weight compatible with that of the mammalian UCN. In the cloacal burse, immunoreactivity was located in the medullary epithelial cells and in the follicular associated and cortico-medullary epithelium. In the thymus, immunoreactivity was located in single epithelial cells. Double labelling immunofluorescence studies showed that UCN immunoreactivity completely colocalised with cytokeratin immunoreactivity in both the thymus and cloacal burse. Statistically significant differences in the percentage of UCN immunoreactivity were observed between different age periods in the cloacal burse. The results suggest that, in birds, urocortin has an important role in regulating the function of the immune system.

Urocortin-like immunoreactivity in the primary lymphoid organs of the duck (Anas platyrhynchos).

Urocortin (UCN) is a 40 aminoacid peptide which belongs to corticotropin-releasing factor (CRF) family. This family of peptides stimulates the secretion of proopiomelanocortin (POMC)-derived peptides, adrenocorticotropic hormone (ACTH), β-endorphin and melanocyte-stimulating hormone (MSH) in the pituitary gland. In the present study, using Western blotting and immunohistochemistry, the distribution of UCN in the primary lymphoid organs of the duck was investigated at different ages. In the cloacal burse and thymus, Western blot demonstrated the presence of a peptide having a molecular weight compatible with that of the mammalian UCN. In the cloacal burse, immunoreactivity was located in the medullary epithelial cells and in the follicular associated and corticomedullary epithelium. In the thymus, immunoreactivity was located in single epithelial cells. Double labelling immunofluorescence studies showed that UCN immunoreactivity completely colocalised with cytokeratin immunoreactivity in both the thymus and cloacal burse. Statistically significant differences in the percentage of UCN immunoreactivity were observed between different age periods in the cloacal burse. The results suggest that, in birds, urocortin has an important role in regulating the function of the immune system.

Distribution of urocortin‐like immunoreactivity in the central nervous system of the frog Rana esculenta

Corticotropin-releasing factor (CRF), sauvagine, and urotensin I are all members of the so-called CRF neuropeptide family. Urocortin (Ucn), a 40-amino-acid neuropeptide recently isolated from the rat brain, is the newest member of this family. Until now, the distribution of Ucn in the central nervous system (CNS) has been studied only in placental mammals. We used a polyclonal antiserum against rat Ucn to determine the distribution of Ucn-like immunoreactivity in the CNS of the green frog, Rana esculenta. The great majority of Ucn-immunoreactive perikarya was seen in the anterior preoptic area, ventromedial thalamic nucleus, posterior tuberculum, nucleus of the medial longitudinal fasciculus, and Edinger-Westphal nucleus. Urocortin-immunoreactive nerve cells were also observed in the motor nuclei of the trigeminal and facial nerves and in the hypoglossal nucleus. Immunoreactive fibers were found in the medial and lateral septal nuclei, bed nucleus of the stria terminalis, many of the thalamic and hypothalamic nuclei, mesencephalic tectum, tegmental nuclei, torus semicircularis, and dorsal horn and central field of the spinal cord. Only scattered Ucn-immunoreactive axon terminals were observed in the external zone of the medial eminence. The densest accumulations of Ucn-immunoreactive nerve terminals were seen in the granular layer of the cerebellum and cochlear nuclei. Our results suggest that an ortholog of mammalian Ucn occurs in the CNS of the green frog. The distribution of Ucn-like immunoreactivity in Rana esculenta showed many similarities to the distribution in placental mammals. The distribution of Ucn-like immunoreactivity in the anuran CNS was different from that of CRF and sauvagine, so our results suggest that at least three different lineages of the CRF neuropeptide family occur in the anuran CNS.

Distribution of urocortin in the rat’s gastrointestinal tract and its colocalization with tyrosine hydroxylase

Urocortin (Ucn), a newly identified member of the corticotropin-releasing factor (CRF) family, is not only expressed in the brain, but also abundantly present in the peripheral tissues, especially in the gastrointestinal tract (GI) as determined by radioimmuoassay. In order to determine the precise localization of urocorin in the GI, we mapped the distribution of urocortin-like immunoreactivity (ir) in the GI of the rat using an immunofluorescence histochemical technique. Ucn, both in the brain and the peripheral tissues, is involved in the regulatory control of host-defense mechanism during stress. In order to study the possible involvement of the sympathetic system in the expression of GI urocortin in response to stress, we examined the effect of chemical sympathectomy on urocortin-ir and its colocalization with tyrosine hydroxylase (TH). UCn was expressed in all parietal cells of the stomach, myenteric and submucosal plexuses as well as in cells in Lieberkühn crypts of the small and large intestine. Most of the acid secreting parietal cells contained both Ucn and TH. Chemical sympathectomy did not affect Ucn immunoreactivity of parietal cells.

Increase of urocortin-like immunoreactivity in the supraoptic nucleus of Dahl rats given a high salt diet.

Urocortin-like immunoreactivity (Ucn-LI) in the supraoptic nucleus (SON) of Dahl rats was examined. Dahl salt-sensitive (S) rats fed with a high salt diet developed hypertension. Numbers of Ucn-LI neurons in the SON in Dahl S on a high salt diet were markedly increased, compared with those in Dahl salt-resistant (R) rats on the same. Sporadic Ucn-LI neurons were found in the SON of both Dahl S and R on a normal diet. Numbers of Ucn-LI neurons in the SON of spontaneously hypertensive rat (SHR) and stroke-prone SHR, genetic models of hypertension, and control rats (Sprague-Dawley and Wistar-Kyoto) were similar. These results suggest that Ucn in the SON is associated with salt loading-induced hypertension rather than spontaneous hypertension.

Increase of urocortin-like immunoreactivity in the supraoptic nucleus of rats with aging and hypertensive Dahl rats

Increase of urocortin-like immunoreactivity in the supraoptic nucleus of rats with aging and hypertensive Dahl rats

Increased urocortin-like immunoreactivity in the supraoptic nucleus of rats with aging

Increased urocortin-like immunoreactivity in the supraoptic nucleus of rats with aging

Differential distribution of urocortin- and corticotropin-releasing factor-like immunoreactivities in the rat brain

Urocortin, a novel 40 amino acid neuropeptide, is a member of the corticotropin-releasing factor family. With 45% homology to corticotropin-releasing factor, urocortin binds with similar affinity to the corticotropin-releasing factor-1 and corticotropin-releasing factor-2 receptors and may play a role in modulating many of the same systems as corticotropin-releasing factor. To assess whether urocortin and corticotropin-releasing factor are localized in the same regions of the brain, we compared the distribution of urocortin- and corticotropin-releasing factor-like immunoreactivities in the rat central nervous system. Polyclonal antibodies to rat corticotropin-releasing factor and rat urocortin were generated and utilized to map the distribution of corticotropin-releasing factor- and urocortin-like immunoreactivities throughout the rat forebrain and brainstem. Characterization of the antibodies by radioimmunoassay showed no cross-reactivity with related peptides. Male Sprague–Dawley rats were treated with colchicine for 18–24 h. Following colchicine treatment, the rats were perfused with paraformaldehyde–lysine–periodate fixative and their brains removed. Serial coronal sections were taken throughout the rat brain and processed for either corticotropin-releasing factor- or urocortin-like immunoreactivity. Urocortin-like immunoreactivity shows a discrete localization within several regions including the supraoptic nucleus, the median eminence, Edinger–Westphal nucleus and the sphenoid nucleus. This is in contrast to the more abundant corticotropin-releasing factor-like immunoreactivity. Regions containing high levels of corticotropin-releasing factor immunoreactivity include the lateral septum, paraventricular nucleus of the hypothalamus, median eminence and locus coeruleus. There are a few regions that contain both urocortin-immunoreactive and corticotropin-releasing factor-immunoreactive cells, such as the supraoptic nucleus and the hippocampus. Therefore, urocortin and corticotropin-releasing factor appear to have different distribution patterns which may be indicative of their respective physiological functions.

Regional Distribution of Urocortin-like Immunoreactivity and Expression of Urocortin mRNA in the Human Brain

Regional distribution of urocortin-like immunoreactivity (UCN-LI) in the human brain was studied by radioimmunoassay and was compared with that of corticotropin-releasing hormone (CRH). In addition, the expression of UCN mRNA was examined by reverse transcriptase-polymerase chain reaction (RT-PCR) method. UCN-LI was detected in every region of brain examined, including hypothalamus, pons, cerebral cortex, and cerebellum. The concentrations of UCN-LI in the human brain were approximately 3 pmol/g wet weight in any brain region, and no marked regional difference was noted. On the other hand, the highest concentrations of CRH-LI were found in the frontal cortex, temporal cortex, and hypothalamus and the lowest in the pons. Reverse phase high-performance liquid chromatography of the UCN-LI in the human brain extract showed two immunoreactive peaks; one peak eluting earlier and one in the position of synthetic human UCN. RT-PCR showed that UCN mRNA was expressed in every region of brain examined. These findings indicated that UCN and UCN mRNA were widely expressed in the human brain.

Distribution of urocortin-like immunoreactivity in the central nervous system of the rat.

Urocortin was recently cloned from the rat midbrain. Urocortin is a member of the corticotropin releasing factor (CRF) peptide family and shows 45% sequence identity to CRF and 63% sequence identity to urotensin. It binds with a high affinity to CRF1 and CRF2 receptors, resulting in the stimulation of their adenylate cyclase activity. We used a polyclonal antibody against rat urocortin to define the distribution of urocortin-like immunoreactivity in the rat central nervous system. Several immunostained cell bodies were found in the supraoptic, paraventricular, and ventromedial hypothalamic nuclei. A large number of neurons with urocortin-like immunoreactivity were seen in the dorsolateral tegmental nucleus, in the linear and dorsal raphe nuclei, and in the substantia nigra. The most abundant immunoreactive (ir) perikarya were found in the Edinger-Westphal nucleus. Some neurons showed immunoreactivity in the interstitial nucleus of Cajal, the nucleus of Darkeschewitsch, and the periaqueductal gray. A dense immunoreactive fiber network was found in the lateral septal area. Some faintly stained axon terminals were observed among urocortin-ir perikarya in the supraoptic and paraventricular nuclei, in the central and periaqueductal gray, and in the Edinger-Westphal nucleus. No fibers with urocortin-ir were seen in the median eminence or the posterior pituitary. The distribution of urocortin-ir overlapped with the expression of the mRNA for the CRF2 receptor in several brain areas. These data support the hypothesis that this peptide is the endogenous ligand for the CRF2 receptor. Urocortin has been implicated in various endocrine responses, such as blood pressure regulation, as well as in higher cognitive functions.

Urocortin-like immunoreactivity in the substantia nigra, ventral tegmental area and Edinger–Westphal nucleus of rat

Neurons showing intense urocortin-like immunoreactivity (Ucn-IR) were found immunohistochemically in the substantia nigra pars compacta (SNc) and ventral tegmental area (VTA), as well as in Edinger–Westphal nucleus (E–W), in the rat brain. Almost all Ucn-immunoreactive neuronal cell bodies in the SNc and VTA showed immunoreactivity for tyrosine hydroxylase. Injection of a retrograde tracer, colloidal gold-labeled WGAapoHRP, into the cervical spinal cord resulted in labeling of some E–W neurons with Ucn-IR. These findings suggest that Ucn is located in the cell bodies of dopaminergic neurons, as well as in the cell bodies of E–W neurons sending axons to the spinal cord.

Increase of urocortin-like immunoreactivity in the rat supraoptic nucleus after dehydration but not food deprivation

The effects of dehydration and food deprivation on urocortin-like immunoreactivity (Ucn-IR) in the rat supraoptic nucleus (SON) and the paraventricular nucleus (PVN) were examined by immunohistochemistry. Water deprivation for 48 h caused a significant increase in the number of Ucn-IR neurons in the SON, compared with control. Ucn-IR fibers and varicosities in the SON and the internal zone of the median eminence (ME) were increased, but a few and faint Ucn-IR neurons and fibers were observed in the PVN. On the other hand, food deprivation for 48 h caused a significant decrease in the number of Ucn-IR neurons in the SON, compared with control. Ucn-IR fibers and varicosities in the SON and the ME were fewer than those in controls. Ucn-IR neurons and fibers in the PVN were not detected after food deprivation. These results suggest that Ucn in the SON may be involved in the central regulation of water balance and nutrient homeostasis.

Increase of urocortin-like immunoreactivity in the rat hypothalamo-neurohypophysial system after salt loading and hypophysectomy

The effect of chronic salt loading on urocortin-like immunoreactivity (Ucn-IR) was investigated in the rat hypothalamo-neurohypophysial system. In control rats a few Ucn-IR neurons were observed scattered throughout the supraoptic nucleus (SON) and few in the paraventricular nucleus (PVN). A small number of Ucn-IR fibers were observed scattered in the median eminence (ME) and the posterior pituitary. However, after 5 days of chronic administration of 2% saline, a marked increase in the number of Ucn-IR perikarya and fibers was observed in the PVN and the SON. Additionally, Ucn-IR varicosities and fibers were found in the internal zone of the ME and in the posterior pituitary. To confirm the findings and examine the possible involvement of anterior pituitary function in synthesis of Ucn, surgical hypophysectomized rats were used. Five days after hypophysectomy, a marked increase in Ucn-IR was observed in the PVN, the SON, and both the internal and the external zone of the ME. These results suggest that Ucn in the hypothalamo-neurohypophysial system may be involved in the regulation of salt balance, and possibly in the stimulation of ACTH release from the anterior pituitary.