Innovative high-resolution microCT imaging of animal brain vasculature.

Analysis of the angioarchitecture and quantification of the conduit vessels and microvasculature is of paramount importance for understanding the physiological and pathological processes within the central nervous system (CNS). Most of the available in vivo imaging methods lack penetration depth and/or resolution. Some ex vivo methods may provide better resolution, but are mainly destructive, as they are designed for imaging the CNS tissues after their removal from the skull or vertebral column. The removal procedure inevitably alters the in situ relations of the investigated structures and damages the dura mater and leptomeninges. µAngiofil, a polymer-based contrast agent, permits a qualitatively novel postmortem microangio-computed tomography (microangioCT) approach with excellent resolution and, therefore, visualization of the smallest brain capillaries. The datasets obtained empower a rather straightforward quantitative analysis of the vascular tree, including the microvasculature. The µAngiofil has an excellent filling capacity as well as a radio-opacity higher than the one of bone tissue, which allows imaging the cerebral microvasculature even within the intact skull or vertebral column. This permits in situ visualization and thus investigation of the dura mater and leptomeningeal layers as well as their blood supply in their original geometry. Moreover, the methodology introduced here permits correlative approaches, i.e., microangioCT followed by classical histology, immunohistochemistry and even electron microscopy. The experimental approach presented here makes use of common desktop microCT scanners, rendering it a promising everyday tool for the evaluation of the (micro)vasculature of the central nervous system in preclinical and basic research.

The Impact of Radiation Energy on Dose Homogeneity and Organ Dose in the Göttingen Minipig Total-Body Irradiation Model.

Animal models of total-body irradiation (TBI) are used to elucidate normal tissue damage and evaluate the efficacy of medical countermeasures (MCM). The accuracy of these TBI models depends on the reproducibility of the radiation dose-response relationship for lethality, which in turn is highly dependent on robust radiation physics and dosimetry. However, the precise levels of radiation each organ absorbs can change dramatically when different photon beam qualities are used, due to the interplay between their penetration and the natural variation of animal sizes and geometries. In this study, we evaluate the effect of varying the radiation energy, namely cobalt-60 (Co-60); of similar penetration to a 4-MV polyenergetic beam), 6 MV and 15 MV, in the absorbed dose delivered by TBI to individual organs of eight Göttingen minipigs of varying weights (10.3-24.1 kg) and dimensions (17.5-25 cm width). The main organs, i.e. heart, lungs, esophagus, stomach, bowels, liver, kidneys and bladder, were contoured by an experienced radiation oncologist, and the volumetric radiation dose distribution was calculated using a commercial treatment planning system commissioned and validated for Co-60, 6-MV and 15-MV teletherapy units. The dose is normalized to the intended prescription at midline in the abdomen. For each animal and each energy, the body and organ dose volume histograms (DVHs) were computed. The results show that more penetrating photon energies produce dose distributions that are systematically and consistently more homogeneous and more uniform, both within individual organs and between different organs, across all animals. Thoracic organs (lungs, heart) received higher dose than prescribed while pelvic organs (bowel, bladder) received less dose than prescribed, due to smaller and wider separations, respectively. While these trends were slightly more pronounced in the smallest animals (10.3 kg, 19 cm abdominal width) and largest animals (>20 kg, ∼25 cm abdominal width), they were observed in all animals, including those in the 9-15 kg range typically used in MCM models. Some organs received an average absorbed dose representing <80% of prescribed dose when Co-60 was used, whereas all organs received average doses of >87% and >93% when 6 and 15 MV were used, respectively. Similarly, average dose to the thoracic organs reached as high as 125% of the intended dose with Co-60, compared to 115% for 15 MV. These results indicate that Co-60 consistently produces less uniform dose distributions in the Göttingen minipig compared to 6 and 15 MV. Moreover, heterogeneity of dose distributions for Co-60 is accentuated by anatomical and geometrical variations across various animals, leading to different absorbed dose delivered to organs for different animals. This difference in absorbed radiation organ doses, likely caused by the lower penetration of Co-60 and 6 MV compared to 15 MV, could potentially lead to different biological outcomes. While the link between the dose distribution and variation of biological outcome in the Göttingen minipig has never been explicitly studied, more pronounced dose heterogeneity within and between organs treated with Co-60 teletherapy units represents an additional confounding factor which can be easily mitigated by using a more penetrating energy.

Ex vivo diffusion-weighted MRI tractography of the Göttingen minipig limbic system.

The limbic system encompasses a collection of brain areas primarily involved in higher cognitive and emotional processing. Altered function in the limbic circuitry may play a major role in various psychiatric disorders. This study aims to provide a high-quality ex vivo diffusion-weighted MRI (DWI) tractographic overview of the Göttingen minipig limbic system pathways, which are currently not well described. This may facilitate future translational large animal studies. The study used previously obtained post-mortem DWI scans in 3 female Göttingen minipigs aging 11-15 months. The tractography performed on the DWI data set was made using a probabilistic algorithm, and regions of interest (ROIs) were defined in accordance with a histological atlas. The investigated pathways included the fornix, mammillothalamic tract, stria terminalis, stria medullaris, habenulo-interpeduncular tract, and cingulum. All the investigated limbic connections could be visualized with a high detail yielding a comprehensive three-dimensional overview, which was emphasized by the inclusion of video material. The minipig limbic system pathways displayed using tractography closely resembled what was previously described in both human studies and neuronal tracing studies from other mammalian species. We encountered well-known inherent methodological challenges of tractography, e.g., partial volume effects and complex white matter regions, which may have contributed to derouted false-positive streamlines and the failure to visualize some of the minor limbic pathway ramifications. This underlines the importance of preexisting anatomical knowledge. Conclusively, we have, for the first time, provided an overview and substantial insight of the Göttingen minipig limbic system.

Anatomical feature of knee joint in Aachen minipig as a novel miniature pig line for experimental research in orthopaedics.

BACKGROUND: The pig is a commonly used large animal model, since pigs share anatomical and physiological similarities with humans. In contrast to other experimental pig lines the Aachen minipig, as a robust novel minipig does not require housing with any barrier. To estimate transferability of results to human conditions, pig lines should be thoroughly characterized. PURPOSE: Therefore, we analyzed the anatomical pecularities of the knee joint of the novel "Aachen minipig" line raised for experimental conditions. METHODS: Eight knee joints of four adult Aachen minipigs were dissected measuring the dimensions of typical landmarks using a digital caliper. Hybrid pig and human knee joints served as controls. Cartilage of the Aachen minipig (trochlear groove, femoral condyles, menisci) were assessed histologically. RESULTS: The Aachen minipig shared its knee joint anatomy with the hybrid pig. In comparison to humans, peculiarities of the pig were demonstrated in the Aachen minipig: the lateral meniscus and the lateral tibial joint surface were significantly longer than the medial counterparts. The fibular head was covered by fibrocartilage and completely integrated into the lateral lower joint surface. The cartilage at the joint areas usually used for cartilage repair studies was in average 0.66±0.04mm thick. The porcine anterior cruciate ligament (ACL) attached with two bundles at the anterior tibial plateau separated from each other by the lateral anterior meniscotibial ligament. Aachen minipig articular and meniscal cartilage presented the typical histoarchitecture. CONCLUSIONS: The Aachen minipig reflects porcine anatomical peculiarities, which should be considered, especially for meniscus and ACL reconstruction.

Spontaneous Age-Related Histopathological Changes in Microminipigs.

Microminipigs have become an attractive animal model for toxicology and pharmacology studies and for human disease models, owing to their manageable size. Although there are numerous reports of spontaneous age-related lesions in mice, rats, dogs, and monkeys, those in minipigs are scarce. In the present study, spontaneous age-related histopathological changes were investigated using 37 microminipigs (20 males and 17 females) that were 6 months to 10 years of age. Abnormal deposits of materials were evident in several animals from 6 years of age, and these deposits included amyloid in the renal medulla, thyroid gland, and adrenal gland, hyaline droplets in glomeruli, and fibrillar inclusions in neurons. Arterial sclerosing changes (intimal thickening, intimal proliferation, and medial mineralization) and proliferative lesions (hyperplasia of hepatocytes, follicular cells, Leydig cells, and uterine endometrial glands) were present at 4 years of age and beyond. Renal adenoma, uterine leiomyoma, and Leydig cell tumor were observed in several microminipigs. Moreover, glomerulosclerosis, renal interstitial fibrosis, thymic involution, and adrenocortical cell vacuolation were common in aging microminipigs. Since knowledge of age-related changes is helpful for pathologists, the basic information obtained in this study will be a useful reference for all future toxicity evaluations in microminipigs.

Histomorphometric evaluation of the Göttingen minipig eye.

OBJECTIVE: The Göttingen minipig is becoming a popular nonrodent animal model in ocular research; however, there is a paucity of literature regarding normative ocular reference data in this breed. We, therefore, investigated the characteristics of the cornea, retina, and sclera in order to establish baseline histomorphometric data in male and female Göttingen minipigs. PROCEDURES: This study utilized paraffin-embedded, Davidson's-fixed, control Göttingen minipig eyes (six males and eight females). Hematoxylin and eosin stained slides of the eyes were scanned via an Aperio slide scanner and analyzed using Aperio ImageScope™. Linear measurements were made of the cornea, retina, and sclera. RESULTS: There were no statistically significant differences between males and females in corneal or scleral measurements or total retinal thickness. There were, however, statistically significant differences between the sexes in the thickness of the ventral peripheral ganglion cell layer [13.15 µm (±3.65) in males; 10.68 µm (±3.37) in females; P = 0.03], ventral peripheral inner plexiform layer [23.47 µm (±2.85) in males; 21.16 µm (±3.62) in females; P = 0.03], ventral central outer plexiform layer [7.97 µm (±2.43) in males; 6.63 µm (±1.73) in females; P = 0.02], and ventral peripheral outer plexiform layer [8.79 µm (±1.82) in males; 11.23 µm (±3.11) in females; P = 0.01]. CONCLUSIONS: This study provides normative histomorphometric reference data for the Göttingen minipig eye. These data will aid researchers in study design and interpretation of findings in Göttingen minipig ocular studies.

Establishment of a Large Animal Model for Eustachian Tube Functional Study in Miniature Pigs.

This study was performed to investigate whether miniature pigs are a suitable animal model for studies of the Eustachian tube (ET). Sixteen Chinese experimental miniature pigs were used in this investigation. Ten animals were used for anatomical and morphometric analyses to obtain qualitative and quantitative information regarding the ET. Three animals were used for histological analysis to determine the fine structure of ET cross-sections. Three animals were used to investigate the feasibility of balloon dilation of the Eustachian tube (BDET). The anatomical study indicated that the pharyngeal orifice and tympanic orifice of the miniature pig ET are located at the posterior end of the nasal lateral wall and anterior wall of the middle ear cavity, respectively. The cartilaginous tube was seen to pass through the whole length of the ET, the length of the cartilaginous part of the ET and the diameter of the isthmus were similar between humans and miniature pigs. The inclination of the ET in miniature pigs was larger than that in humans. The gross histology seemed to be slightly different between miniature pig and human, but the fine structures were essentially the same in both species. BDET experiments verified that the miniature pig model is suitable as a model for clinical operations. The miniature pig ET corresponds very well to that of humans. In addition, the miniature pig ET is suitable as a model for clinical operations. Therefore, the miniature pig is a valid animal model for ET study. Anat Rec, 302:1024-1038, 2019. © 2019 Wiley Periodicals, Inc.

The porcine corticospinal decussation: A combined neuronal tracing and tractography study.

BACKGROUND: Pigs and minipigs are increasingly used as non-primate large animal models for preclinical research on nervous system disorders resulting in motor dysfunction. Knowledge of the minipig pyramidal tract is therefore essential to support such models. AIM AND METHODS: This study used 5 female Göttingen minipigs aging 11-15 months. The Göttingen minipig corticospinal tract was investigated, in the same animals, with in vivo neuronal tracing and with postmortem diffusion weighted MRI tractography to provide a thorough insight in the encephalic distribution of this primary motor pathway and its decussation at the craniocervical junction. RESULTS: The two methods similarly outlined the course of the pyramidal tract from its origin in the motor cortex down through the internal capsule to the craniocervical junction, where both methods displayed an axonal crossover at the pyramid decussation. The degree of crossover was quantified with unbiased stereology, where 81-93% of the traced corticospinal fibers crossed to the contralateral spinal cord. Accordingly, in the upper cervical spinal cord the corticospinal tract is primarily distributed in the contralateral lateral funiculus and in close relation to the gray matter, wherein some direct terminations on large ventral column gray matter neurons could be identified. DISCUSSION: The combination of neuronal tracing and tractography exploited the strengths of the respective methods to gain a better understanding of the encephalic distribution and craniocervical decussation of the Göttingen minipig corticospinal tract. Moreover, a quantification of the crossing fibers was obtained from the tracing data, which was not possible with tractography. Our data indicate that the porcine corticospinal system is quite lateralized down to the investigated upper cervical levels. However, further elucidation of this point will require a full examination of the corticospinal tracing pattern into the caudal spinal cord combined with an analysis of the direct versus indirect termination pattern on the lower motor neurons.

Expression patterns of tight junction proteins in porcine major salivary glands: a comparison study with human and murine glands.

Tight junction (TJ) proteins play a dynamic role in paracellular fluid transport in salivary gland epithelia. Most TJ studies are carried out in mice and rats. However, the morphology of rodent salivary glands differs from that of human glands. This study aimed to compare the histological features and the expression pattern of TJ proteins in porcine salivary glands with those of human and mouse. The results showed that porcine parotid glands were pure serous glands. Submandibular glands (SMGs) were serous acinar cell-predominated mixed glands, whereas sublingual glands were mucous acinar cell-predominated. Human SMGs were mixed glands containing fewer mucous cells than porcine SMGs, whereas the acinar cells of murine SMGs are seromucous. The histological features of the duct system in the porcine and human SMGs were similar and included intercalated, striated and excretory ducts, but the murine SMG contained a specific structure, the granular convoluted tubule. TJ proteins, including claudin-1 to claudin-12, occludin and zonula occludin-1 (ZO-1), were detected in the porcine major salivary glands and human SMGs by RT-PCR; however, claudin-6, claudin-9 and claudin-11 were not detected in the murine SMG. As shown by immunofluorescence, claudin-1, claudin-3, claudin-4, occludin and ZO-1 were distributed in both acinar and ductal cells in the porcine and human SMGs, whereas claudin-1 and claudin-3 were mainly present in acinar cells, and claudin-4 was mainly distributed in ductal cells in the murine SMG. In addition, 3D images showed that the TJ proteins arranged in a honeycomb-like structure on the luminal surface of the ducts, whereas their arrangements in acini were irregular in porcine SMGs. In summary, the expression pattern of TJ proteins in salivary glands is similar between human and miniature pig, which may be a candidate animal for studies on salivary gland TJ function.

Normative Retinal Thicknesses in Common Animal Models of Eye Disease Using Spectral Domain Optical Coherence Tomography.

Purpose This study demonstrates a standardized approach to measuring retinal thickness (RT) using spectral domain optical coherence tomography (SD-OCT) in commonly used animal models of disease and reports a normative data set for future use. Materials and Methods Twenty normal eyes of 4 adult animal models (5 rats, 5 rabbits, 5 canines, and 5 mini-pigs) were used. Manual measurements were made on the commercially available Heidelberg Spectralis™ SD-OCT to determine the total, inner, and outer retinal thickness (RT) at fixed distances from the optic nerve head (ONH) (1, 2, 3, 4, 5, and 6 mm away) in order to control for normal variation in retinal thickness. Analysis of variance (ANOVA) with P value <0.05 indicated statistical significance. Results Total RT significantly decreased with increasing distance from the ONH for the canine, mini-pig, and rabbit vascular models. Inner RT significantly decreased for the canine, mini-pig, rabbit vascular, and rabbit avascular models; and outer RT significantly decreased for only the canine model. Among the animal models, RT at similar distances from the ONH were significantly different for total, inner, and outer RT. Conclusion There are significant differences in the total, inner, and outer RT of normal canine, mini-pig, rabbit, and rat retinas with SD-OCT using a standardized approach. These measurements provide a normative reference for future studies and illustrate a standardized method of assessing RT.

A reproducible swine model of proximal descending thoracic aortic aneurysm created with intra-adventitial application of elastase.

OBJECTIVE: Animal models are required to explore the mechanisms of and therapy for proximal descending thoracic aortic aneurysm (TAA). This study aimed to establish a reproducible swine model of proximal descending TAA that can further explain the occurrence and progression of proximal descending TAA. METHODS: Eighteen Chinese Wuzhishan miniature pigs (30.32 ± 1.34 kg) were randomized into the elastase group (n = 12) and the control group (n = 6). The elastase group received intra-adventitial injections of elastase (5 mL, 20 mg/mL), and the control group received injections of physiologic saline solution. A 4-cm descending thoracic aortic segment proximal to the left subclavian artery was isolated. The distance between the left subclavian artery and the injection starting point of the descending thoracic aorta was 0.5 cm. Elastic protease was circumferentially injected intra-adventitially into the isolated segment of the aortic wall in the elastase group by a handmade bent syringe. The length of the elastic protease injection was 2 cm. An average of 12 injection points were distributed in this 2-cm aortic segment. Each injection point used about 0.4 mL of elastic protease. The distance between two injection points was about 1.5 cm. All animals underwent digital subtraction angiography preoperatively and 3 weeks after operation. Three weeks after TAA induction, aortas were harvested for biochemical and histologic measurements. RESULTS: All animals in the elastase group developed TAAs. No aneurysms were observed in the control group. The distance between the left subclavian artery and the TAA was 8.00 ± 4.19 mm. Preoperative and postoperative aortic diameters of the elastase group were 15.42 ± 0.43 mm and 24.53 ± 1.41 mm, respectively (P < .0001). Preoperative and postoperative aortic diameters of the control group were 15.31 ± 0.33 mm and 15.57 ± 0.40 mm, respectively (P = .5211). The changes of aortic structure and composition included reduction of smooth muscle cells and degradation of elastic fibers. Levels of matrix metalloproteinases 2 and 9 were increased in TAA tissue. CONCLUSIONS: This study established a reproducible large animal model of proximal descending TAA. This model has the same biochemical characteristics as human aneurysms in the aspects of aortic expansion, aortic middle-level degeneration, and changes in the levels of matrix metalloproteinases and provides a platform for further study.

Normal Developmental and Estrous Cycle-dependent Histological Features of the Female Reproductive Organs in Microminipigs.

The microminipig has become an increasingly attractive animal model for various experimental practices because of its manageable size; however, studies of the histological features of the female reproductive organs in microminipigs are limited. The present study investigates the sexual development of the reproductive organs and the cyclical changes during the estrous cycle in female microminipigs. The ovaries, oviducts, uteri, and vaginal tissues from 33 animals aged 0 to 26 months were utilized in this study. By evaluating the large tertiary follicles, corpora lutea, and the regressing corpora lutea, we estimated that female microminipigs reached puberty at approximately 5 months of age and sexual maturity at 8 months of age. The appearance of the follicles and corpora lutea in the ovaries, as well as the epithelium in other reproductive organs, was synchronized with each phase of the estrous cycle and was identical to that in common domestic pigs. In addition, several spontaneous findings were observed, including mesonephric duct remnants adjacent to oviducts and mineralization in ovaries. Understanding the normal histology of the reproductive organs in microminipigs is crucial for advancing pathological evaluations for future toxicological studies.

Body surface area measurement in juvenile miniature pigs using a computed tomography scanner.

The use of miniature pigs in non-clinical studies for medical drugs or devices has gradually been increasing in recent years. It is anticipated that the use of juvenile miniature pigs in laboratory practice will also increase. Therefore, it is important to investigate various parameters of juvenile miniature pigs. The body surface area (BSA) of an organism is one of the important parameters for evaluating physiological functions. In drug development, normalization by BSA is an appropriate method for extrapolating doses between species. The BSA of animals has generally been estimated by multiplying the k value by 2/3 of the power of the body weight (BW) (Meeh's formula). To our knowledge, the BSA of juvenile miniature pigs has not as yet been reported. In this study, we measured the BSA of 13 miniature pigs less than 1 month old, using a computed tomography scanner and 3-dimensional analysis software. The measurement results showed the BSAs of these 13 juvenile miniature pigs to be in the range of 386 to 1,672 cm2(working BW range: 278 to 3,200 g). After BSA determination, the k values were calculated from the BSA and the BW. The mean calculated k value was 8.58. We advocate using Meeh's formula, as follows, for estimating the BSA of juvenile miniature pigs less than 1 month old (before weaning): BSA (cm2)=8.58 × BW (g)2/3.

Histological and anatomical structure of the nasal cavity of Bama minipigs.

OBJECTIVE: The nasal mucosa is equipped with abundant lymphatic tissues, serving as the first line of defense against invasion by microorganisms. In this study, we characterized the features of the nasal mucosa of Bama minipigs (Sus scrofa domestica) via histological analysis. METHODS: Five cross sections (I, II, III, IV, and V) were obtained from the distal end of the nasal cavity toward the pharynx (along the cavity axis) and examined. Specifically, CD3+ T cells, immunoglobulin A (IgA)+ cells, and M cells were detected by immunohistochemistry, while dendritic cells (DCs) were detected by immunofluorescence. The distribution of goblet cells was determined by periodic acid-Schiff (PAS) staining. RESULTS: The nasal cavity of Bama minipigs can be divided into three parts: the regio vestibularis (I, II), regio respiratoria (III, IV), and regio olfactoria (V). Lymphoid tissue was present at random locations in the nasal cavity. Abundant lymphoid tissue was located in the roof of the nasopharyngeal meatus and was continuous with the lymphoid tissue of the pharynx. The distribution of CD3+ T cells, IgA+ cells, M cells, and DCs increased distally in the nasal cavity. CONCLUSIONS: The present work comprises a histological study of the nasal cavity of Bama minipigs, and will be beneficial for understanding the mechanisms of immunity in these animals after nasal vaccination.

A Comprehensive Study of Palate Development in Miniature Pig.

Palate development is an important morphogenetic event in facial development, including the fusion of the lateral and medial nasal portions of the frontonasal process and maxilla. Derailments of any of these events may result in cleft palate, the most frequent congenital craniofacial abnormality. Recent research has shown that the microanatomy of the miniature pig oral maxillofacial region is quite similar to that of humans, and the use of miniature pigs as a large animal model for dental and orofacial research is increasing. Little information is available, however, about the development of the miniature pig palate. Here, using histological and ultrastructural methods, we describe the developmental stages of the palate in miniature pigs. Sections from E26, E30, E35, E40, E45, and E50 embryos were stained with hematoxylin-eosin, and selected specimens were also processed for electron microscopy. The development of the miniature pig palate can be divided into four stages: growth of the bilateral palatal shelves alongside the tongue at E30; elevation of the horizontal position above the tongue at E35; establishment of bilateral shelf contact at the midline from E35-50; and a final fusion step at E50, similar to the mouse and human. The histological characteristics of the miniature pig palate at different developmental stages were synchronously verified at the ultrastructural level. Our study provides a piece of first-hand data regarding palate morphological organogenesis in the miniature pig and a foundation for further research with this model to explore mechanisms of cleft palate development. Anat Rec, 300:1409-1419, 2017. © 2017 Wiley Periodicals, Inc.

Computed tomography-based survey of the vascular anatomy of the juvenile Göttingen minipig.

Over the past 50 years, image-guided procedures have been established for a wide range of applications. The development and clinical translation of new treatment regimens necessitate the availability of suitable animal models. The juvenile Göttingen minipig presents a favourable profile as a model for human infants. However, no information can be found regarding the vascular system of juvenile minipigs in the literature. Such information is imperative for planning the accessibility of target structures by catheterization. We present here a complete mapping of the arterial system of the juvenile minipig based on contrast-enhanced computed tomography. Four female animals weighing 6.13 ± 0.72 kg were used for the analyses. Imaging was performed under anaesthesia, and the measurement of the vascular structures was performed independently by four investigators. Our dataset forms a basis for future interventional studies in juvenile minipigs, and enables planning and refinement of future experiments according to the 3R (replacement, reduction and refinement) principles of animal research.

The importance of minipigs in dermal safety assessment: an overview.

The use of miniature swine as a non-rodent species in safety assessment has continued to expand for over a decade and their use has become routine, particularly in pharmacology as a model for human integumentary diseases. Translational preclinical swine study data are now favorably compared and contrasted to human data, and miniature swine models provide important information in dermal safety assessment and skin pharmacology. For example, the miniature swine model has been well-accepted for cutaneous absorption and toxicity studies due to swine integument being morphologically and functionally similar to human skin. Subsequently, this model is important to dermal drug development programs, and it is the animal model of choice for assessment of dermal absorption, local tolerance and systemic toxicity following dermal exposures. In conclusion, the miniature swine model has an important role to play in the safety assessment of pharmaceutical products and in multiple aspects of human dermal drug development.

Cholinergic profiles in the Goettingen miniature pig (Sus scrofa domesticus) brain.

Central cholinergic structures within the brain of the even-toed hoofed Goettingen miniature domestic pig (Sus scrofa domesticus) were evaluated by immunohistochemical visualization of choline acetyltransferase (ChAT) and the low-affinity neurotrophin receptor, p75NTR . ChAT-immunoreactive (-ir) perikarya were seen in the olfactory tubercle, striatum, medial septal nucleus, vertical and horizontal limbs of the diagonal band of Broca, and the nucleus basalis of Meynert, medial habenular nucleus, zona incerta, neurosecretory arcuate nucleus, cranial motor nuclei III and IV, Edinger-Westphal nucleus, parabigeminal nucleus, pedunculopontine nucleus, and laterodorsal tegmental nucleus. Cholinergic ChAT-ir neurons were also found within transitional cortical areas (insular, cingulate, and piriform cortices) and hippocampus proper. ChAT-ir fibers were seen throughout the dentate gyrus and hippocampus, in the mediodorsal, laterodorsal, anteroventral, and parateanial thalamic nuclei, the fasciculus retroflexus of Meynert, basolateral and basomedial amygdaloid nuclei, anterior pretectal and interpeduncular nuclei, as well as select laminae of the superior colliculus. Double immunofluorescence demonstrated that virtually all ChAT-ir basal forebrain neurons were also p75NTR -positive. The present findings indicate that the central cholinergic system in the miniature pig is similar to other mammalian species. Therefore, the miniature pig may be an appropriate animal model for preclinical studies of neurodegenerative diseases where the cholinergic system is compromised. J. Comp. Neurol. 525:553-573, 2017. © 2016 Wiley Periodicals, Inc.

Histological Development of Male Reproductive Organs in Microminipigs.

Microminipigs are becoming increasingly attractive alternatives for various experimental applications, such as general toxicology studies, owing to their manageable size. However, there are limited studies on the male reproductive organs of microminipigs, particularly on the histological aspects of sexual maturity. To clarify the development of male reproductive organs, 35 male microminipigs, aged 0 to 12 months, were used in this study. Histological and histomorphological evaluation was performed based on spermatogenic development, measurement of tubular structure in testes and epididymides, and histological progress of accessory glands. In addition, spontaneous testicular changes were quantitatively assessed. Histologically, male microminipigs sexually matured around 4.5 months of age, when spermatogenesis in testes and structural development in genital organs were completed. Spontaneous testicular changes occurred in all the animals investigated. Multinucleated giant cell was most commonly observed, followed by hypospermatogenesis and tubular atrophy/hypoplasia. However, the number of affected tubules was less than 1% in testes after 4.5 months of age, suggesting that the influence of these changes on evaluation of toxicity studies may be minimal. It is preferable to use sexually mature animals in toxicology studies; therefore, the information obtained by the present study will be helpful for future toxicity evaluations in microminipigs.