Localization of aquaporin water channels in the airway of the musk shrew (Suncus murinus) and the rat

Aquaporins (AQPs) constitute a family of water channels that facilitate membrane water permeability in various tissues of animals. In this study, we compared the expression and localization of AQPs in the respiratory system of the musk shrew (Suncus murinus), which is an insectivore, and the rat by immunohistochemical methods. In both the musk shrew and the rat, AQP1 was expressed throughout the airway in endothelial cells of subepithelial blood vessels and in nasal submucosal fibroblasts. AQP3 and AQP4 were detected in neither the epithelium nor the subepithelial layer of the musk shrew airway, but were abundant in the rat airway epithelium. Musk shrew AQP5 was distributed in the superficial epithelial cells facing the airspaces and in submucosal glandular cells, but, unlike in the rat, not in lung alveolar cells. Additionally, the expression patterns of AQP4 and AQP5 of the musk shrew were partly similar to those of the human previously reported, absence of AQP4 and presence of AQP5 in the upper airway. The expression differences of AQPs between species in the airway indicate that the physiological importance of each AQP may be different in each species.     

Expression of cysteine sulfinate decarboxylase mRNA in rat mammary gland

Our previous report demonstrated that high concentration of taurine is present in rat milk for the first few days of lactation and plays an important role in the body growth of rat pups. In the present study, gene expression of rate-limiting enzyme for taurine biosynthesis, cysteine sulfinate decarboxylase (CSD) were examined in rat mammary gland. By RT-PCR, CSD mRNA was found to be expressed in rat mammary gland like that in the liver. The expression level of CSD mRNA in the mammary gland was higher in the earlier lactational stage (days 1 and 6 of lactation) than that in the later lactational stage (day 14). CSD mRNA expression in the mammary gland of non-pregnant rats was only a trace level. By in situ hybridization analysis, CSD mRNA was demonstrated in the epithelial cells of the mammary gland. These results suggest that high concentrations of taurine in the milk are at least partially resulted from de novo synthesis of taurine in mammary gland epithelial cells and that the expression pattern of CSD mRNA may be responsible for the changes in taurine levels in the milk during a lactational period.     

Expression of leukemia inhibitory factor and leukemia inhibitory factor receptor in the canine pituitary gland and corticotrope adenomas

Leukemia inhibitory factor (LIF) is a pleiotropic cytokine of the IL-6 family that activates the hypothalamic-pituitary-adrenal axis and promotes corticotrope cell differentiation during development. The aim of this study was to investigate the expression of LIF and its receptor (LIFR) in the canine pituitary gland and in corticotrope adenomas, and to perform a mutation analysis of LIFR. Using immunohistochemistry, immunofluorescence, and quantitative expression analysis, LIF and LIFR expression were studied in pituitary glands of control dogs and in specimens of corticotrope adenoma tissue collected through hypophysectomy in dogs with pituitary-dependent hypercortisolism (PDH, Cushing's disease). Using sequence analysis, cDNA was screened for mutations in the LIFR. In the control pituitary tissues and corticotrope adenomas, there was a low magnitude of LIF expression. The LIFR, however, was highly expressed and co-localized with ACTH_1-24 expression. Cytoplasmatic immunoreactivity of LIFR was preserved in corticotrope adenomas and adjacent nontumorous cells of pars intermedia. No mutation was found on mutation analysis of the complete LIFR cDNA. Surprisingly, nuclear to perinuclear immunoreactivity for LIFR was present in nontumorous pituitary cells of the pars distalis in 10 of 12 tissue specimens from PDH dogs. These data show that LIFR is highly co-expressed with adrenocorticotropic hormone (ACTH) and α-melanocyte-stimulating hormone (α-MSH) in the canine pituitary gland and in corticotrope adenomas. Nuclear immunoreactivity for LIFR in nontumorous cells of the pars distalis may indicate the presence of a corticotrope adenoma.     

Expression of aquaporin 4 in the chicken ovary in relation to follicle development

In the mammalian ovary, aquaporins (AQPs) are thought to be involved in the regulation of fluid transport within the follicular wall and antrum formation. Data concerning the AQPs in the avian ovary is very limited. Therefore, the present study was designed to examine whether the AQP4 is present in the chicken ovary, and if so, what is its distribution in the ovarian compartment of the laying hen. Localization of AQP4 in the ovarian follicles at different stage of development was also investigated. After decapitation of hens the stroma with primordial follicles and white (1–4 mm), yellowish (4–8 mm), small yellow and the three largest yellow pre‐ovulatory follicles F3‐F1 (F3 < F2 < F1; 20–36 mm) were isolated from the ovary. The granulosa and theca layers were separated from the pre‐ovulatory follicles. The AQP4 mRNA and protein were detected in all examined ovarian compartments by the real‐time PCR and Western blot analyses, respectively. The relative expression of AQP4 was depended on follicular size and the layer of follicular wall. It was the lowest in the granulosa layer of pre‐ovulatory follicles and the highest in the ovarian stroma as well as white and yellowish follicles. Along with approaching of the largest follicle to ovulation the gradual decrease in AQP4 protein level in the granulosa layer was observed. Immunoreactivity for AQP4 was present in the granulosa and theca cells (theca interna ≥ theca externa > granulosa). The obtained results suggest that AQP4 may take part in the regulation of water transport required for follicle development in the chicken ovary.     

Mensuration of the rabbit pituitary gland from computed tomography

The aim of this retrospective reference interval observational study was to determine the mensuration of the pituitary gland (hypophysis cerebri) by analyzing CT studies in rabbits without clinical evidence of pituitary disease or central neurologic signs. Though diseases of the rabbit pituitary gland are uncommon, the pituitary gland is essential in regulation of the rabbit's endocrine system, as in other species. Currently, there are minimal published studies that detail the rabbit head anatomy on cross‐sectional imaging, and even less specifically examining the pituitary gland. The pituitary gland was measured by one observer at a single time point from transverse and reconstructed sagittal CT images in a soft‐tissue algorithm in 62 rabbits for a total of 66 rabbit head CT studies. The rabbits ranged from 0.84 to 14 years in age (mean ± SD: 5.46 ± 3.05 years) and 0.92 to 4.95 kg in weight (2.21 ± 0.83 kg). Linear pituitary measurements were performed using electronic calipers. The mean ± SD pituitary height was 4.22 ± 0.57 mm, width was 4.48 ± 0.71 mm, and length was 6.02 ± 0.70 mm. The pituitary gland height‐to‐brain area ratio was 1.10 ± 0.16 mm^?1, which is much higher than the values reported in normal dogs and cats. The age, weight, and sex of the rabbits were not found to have a significant impact on pituitary gland mensuration. These measurements could be useful as a reference range for future rabbit head CT studies and to rule out pituitary enlargement or disease when evaluating rabbit pituitary glands.     

Dynamic computed tomographic evaluation of the pituitary gland in healthy dogs

OBJECTIVE: To determine the contrast enhancement pattern of the pituitary gland in healthy dogs via dynamic computed tomography (CT). ANIMALS: 17 dogs. PROCEDURE: With each dog in sternal recumbency, transverse CT scans were made perpendicular to the skull base from the rostral clinoid processes to the dorsum sellae. At the position of the image that contained the largest cross section of the pituitary gland, a series of 9 to 11 scans was made during and after IV injection of contrast medium (dynamic CT scans). The contrast enhancement pattern of the pituitary gland and surrounding arteries was assessed visually and by use of time-density curves. RESULTS: After strong enhancement of the maxillary arteries, the intracavernous parts of the internal carotid arteries, and the communicating arteries of the arterial cerebral circle, there was a strong enhancement of the central part of the pituitary gland followed by enhancement of its peripheral part. On the last images of the dynamic series of the pituitary gland, the central part was hypodense, compared with the peripheral part. Time-density curves confirmed an early, strong enhancement of the central part and a delayed, less strong enhancement of the peripheral part of the gland. CONCLUSIONS AND CLINICAL RELEVANCE: The difference in enhancement between the central and peripheral parts of the pituitary gland was attributable to a difference in vascularization of the neurohypophysis and adenohypophysis, respectively. Distortion or disappearance of the strong central enhancement (pituitary flush) may be used for the detection and localization of pituitary abnormalities in the adenohypophysis.     

Immunohistochemical localization of S-100 protein in the pig pituitary gland

Immunohistochemical localization of S-100 protein was studied in anterior, intermediate and posterior lobe of the pig pituitary gland. Two immunopositive cells for S-100 protein were identified: the folliculo-stellate cells (FSc) in the glandular lobes and the pituicytes in the neural lobe. In the anterior lobe, immunoreactive folliculo-stellate cells were scattered among secretory cells. In the area where the secretory cells form strands and follicle-like groups the positive cells were concentrated in groups. In the intermediate lobe, S-100 protein-positive cells were located sparsely among secretory cells and next to secretory follicles and the pituitary cleft. These FSc were more voluminous and displayed fewer cytoplasmic processes. In the neurohypophysis, positive reaction for S-100 protein was seen in the pituicytes. These cells were distributed singly or concentrated in groups. The distribution, and morphologic characteristics of the FSc in the glandular lobes and the pituicytes in the neural lobe in the pig indicate different origin of both S-100 protein-positive cells.     

Development of two surgical approaches to the pituitary gland in the Horse

Background: Current treatment of equine pituitary pars intermedia dysfunction (PPID) requires daily oral medication. Minimally invasive surgical palliation of this condition is appealing as a single treatment to alleviate the clinical signs of disease, dramatically improving the welfare of the horse.

Objective: To develop a surgical approach to the equine pituitary gland, for subsequent treatment of PPID.

Study design: A cadaver study to develop methodology and a terminal procedure under anaesthesia in the most promising techniques.

Animals and methods: Four surgical approaches to the pituitary gland were investigated in cadaver animals. A ventral trans-basispheniodal osteotomy and a minimally invasive intravenous approach via the ventral cavernous sinus progressed to live horse trials.

Results: Technical complications prevented the myeloscopic and trans-sphenopalatine sinus techniques from being successful. The ventral basisphenoidal osteotomy was repeatable and has potential if an intra-operative imaging guidance system could be employed. The minimally invasive approach was repeatable, atraumatic and relatively inexpensive.

Conclusions: A minimally invasive surgical approach to the equine pituitary gland is possible and allows for needle placement within the target tissue. More work is necessary to determine what that treatment might be, but repeatable access to the gland has been obtained, which is a promising step.     

Identification and characterisation of side population cells in the canine pituitary gland

To date, stem/progenitor cells have not been identified in the canine pituitary gland. Cells that efficiently exclude the vital dye Hoechst 33342 can be visualised and identified using fluorescence activated cell sorting (FACS) as a 'side population' (SP), distinct from the main population (MP). Such SPs have been identified in several tissues and display stem/progenitor cell characteristics. In this study, a small SP (1.3%, n=6) was detected in the anterior pituitary glands of healthy dogs. Quantitative PCR indicated significantly higher expression of CD34 and Thy1 in this SP, but no differences in the expression of CD133, Bmi-1, Axin2 or Shh. Pro-opiomelanocortin (POMC) and Lhx3 expression were significantly higher in the MP than in the SP, but no differences in the expression of Tpit, GH or PRL were found. The study demonstrated the existence of an SP of cells in the normal canine pituitary gland, encompassing cells with stem cell characteristics and without POMC expression.     

Dynamic computed tomography of the pituitary gland in dogs with pituitary-dependent hyperadrenocorticism

Dynamic computed tomography (CT) of the pituitary gland was performed in 55 dogs with pituitary-dependent hyperadrenocorticism (PDH) that underwent transsphenoidal hypophysectomy. On routine contrast-enhanced CT images, microadenomas of the pituitary gland often are indistinguishable from nontumorous pituitary tissue because of isoattenuation. Dynamic CT may allow visualization of these adenomas. The changes in the contrast-enhancement pattern of the pituitary during dynamic CT in 55 dogs with PDH were correlated with surgical and histopathologic findings. In 36 dogs, dynamic CT identified distinct contrast enhancement of the neurohypophysis (pituitary flush). In 24 dogs, this pituitary flush was displaced, which indicated the presence of an adenoma. This observation was confirmed surgically and histopathologically in 18 of the 24 dogs. In 19 dogs, there was a diffusely abnormal contrast-enhancement pattern. CT findings agreed with surgical findings in 13 of these dogs and with histopathologic findings in all 19 dogs. It is concluded that a dynamic series of scans should be included in the CT protocol of the pituitary gland in dogs with PDH because it allows for identification of an adenoma or a diffusely abnormal pituitary gland.     

The effect of estrogen on phosphorylation of prolactin in the mouse pituitary gland

Several studies have indicated that prolactin (PRL) assumes oligomeric, proteolytically cleaved, phosphorylated and glycosylated forms. Phosphorylated PRL (PPRL) is considered to be the most important posttranslationally modified form in the rat. In the present study, we examined whether or not PRL is present in the mouse pituitary gland in the phosphorylated form. Mouse pituitary PRL was digested with acid phosphatase, resolved by two-dimensional gel electrophoresis, stained with Coomassie brilliant blue, and then immunoblotted against the anti-PRL, anti-phosphoserine and anti-phosphothreonine antibodies. We also examined whether PRL is phosphorylated by protein kinases and semi-quantified the ratios of PPRL to PRL in the pituitary gland. The results indicated that three types of PRL are present in the pituitary glands of both male and female mice. One was non-phosphorylated (isoform 1), and the other two were immunoreactive to anti-phosphoserine (isoform 2) and/or anti-phosphothreonine (isoform 3) antibodies. The ratio between isoforms 2 and 1 of the 30-day-old female mice was higher than that of the 20-day-old female mice. However, the ratios among the three isoforms in the male pituitary glands did not differ with age. The ratio of PPRL to isoform 1 was obviously reduced after ovariectomy (OVX), and it recovered with estrogen replacement. These results suggest that estrogen influences PRL phosphorylation in female mice.     

Expression and localization of tight junction-related proteins in adult rat pituitary stem/progenitor cell niches

Pituitary endocrine cells are supplied by Sox2-expressing stem/progenitor cells in the anterior lobe of the adult pituitary gland. These SOX2-positive cells are maintained in two types of microenvironments (niches): the marginal cell layer (MCL)-niche and the parenchymal-niche. Recently, we isolated dense SOX2-positive cell clusters from the parenchymal-niche by taking advantage of their resistance to protease treatment as parenchymal stem/progenitor cell (PS)-clusters. In the present study, by analyzing these isolated PS-clusters, we attempted to identify novel structural characteristics of pituitary stem/progenitor cell niches. Quantitative real-time PCR showed that tight junction-related genes were distinctly expressed in the isolated PS-clusters. Immunocytostaining showed that the tight junction molecules, ZO-1 and occludin, were localized in the apical membrane facing the pseudo-follicle-like structure of the isolated PS-clusters regardless of the expression of S100β, which distinguishes the sub-population of SOX2-positive cells. Furthermore, immunohistochemistry of the pituitary glands of adult rats clearly demonstrated that ZO-1 and occludin were densely present in the parenchymal-niche encircling the pseudo-follicle, while they were observed in the apical membrane in the MCL-niche facing the residual lumen. Collectively, these tight junction-related proteins might be involved in the architecture and maintenance of the plasticity of pituitary stem/progenitor cell niches.     

Thin-slice three-dimensional gradient-echo magnetic resonance imaging of the pituitary gland in healthy dogs

OBJECTIVE: To evaluate thin-slice 3-dimensional gradient-echo (GE) magnetic resonance imaging (MRI) of the pituitary gland in healthy dogs. ANIMALS: 11 healthy dogs. PROCEDURES: By use of a 0.2-Tesla open magnet, MRI of the skull was performed with T1-weighted GE sequences and various protocols with variations in imaging plane, slice thickness, and flip angle before and after administration of contrast medium; multiplanar reconstructions were made. The pituitary region was subjectively assessed, and its dimensions were measured. Image quality was determined by calculation of contrast-to-noise and signal-to-noise ratios. RESULTS: Best-detailed images were obtained with a T1-weighted GE sequence with 1-mm slice thickness and 30 degrees flip angle before and after administration of contrast medium. Images with flip angles > 50 degrees were of poor quality. Quality of multiplanar reconstruction images with 1-mm slices was better than with 2-mm slices. The bright signal was best seen without contrast medium. With contrast medium, the dorsal border of the pituitary gland was clearly delineated, but lateral borders were more difficult to discern. CONCLUSIONS AND CLINICAL RELEVANCE: MRI of the canine pituitary gland with a 0.2-Tesla open magnet should include a T1-weighted GE sequence with 1-mm slice thickness and flip angle of 30 degrees before and after administration of contrast medium. The neurohypophysis was best visualized without contrast medium. The MRI examination permitted differentiation between the pituitary gland and surrounding structures.     

Differential expression of myostatin and its receptor in the porcine anterior pituitary gland

Myostatin (MSTN), known as growth and differentiation factor 8 (GDF-8), is a member of the transforming growth factor β (TGF-β) superfamily that negatively regulates skeletal muscle mass. Myostatin binds with high affinity to the receptor serine threonine kinase activin receptor type IIB (ActRIIB). Activins that also belong to the TGF-β superfamily, stimulate follicle-stimulating hormone production in gonadotrophs and suppress growth hormone and adrenocorticotropic hormone production in somatotrophs and corticotrophs, respectively. The aim of the present paper was therefore to clarify the endocrine action of MSTN in adenohypophysis. The present study details the expression and cellular localization of MSTN and ActRIIB in porcine anterior pituitary gland. The mRNA of MSTN and ActRIIB was consistently expressed in RT-PCR. Immunohistochemistry of MSTN and specific hormones showed that MSTN localized in thyrotrophs and gonadotrophs, in which most of the MSTN immunoreactive cells were identified as thyrotrophs. The immunostaining of ActRIIB was restricted to corticotrophs. These results indicate that MSTN was mainly produced in thyrotrophs and its receptor, ActRIIB, was restrictively contained in corticotrophs. Interestingly, thyrotrophs immunoreactive for MSTN were frequently close to corticotrophs immunoreactive for ActRIIB. The present study suggests that MSTN from thyrotrophs may regulate corticotroph function as a paracrine mediator among the porcine anterior pituitary cells.