Paternal phylogeography and genetic diversity of East Asian goats.

This study was a first analysis of paternal genetic diversity for extensive Asian domestic goats using SRY gene sequences. Sequencing comparison of the SRY 3'-untranslated region among 210 Asian goats revealed four haplotypes (Y1A, Y1B, Y2A and Y2B) derived from four variable sites including a novel substitution detected in this study. In Asian goats, the predominant haplotype was Y1A (62%) and second most common was Y2B (30%). Interestingly, the Y2B was a unique East Asian Y chromosomal variant, which differentiates eastern and western Eurasian goats. The SRY geographic distribution in Myanmar and Cambodia indicated predominant the haplotype Y1A in plains areas and a high frequency of Y2B in mountain areas. The results suggest recent genetic infiltration of modern breeds into South-East Asian goats and an ancestral SRY Y2B haplotype in Asian native goats.

Molecular phylogeography and genetic diversity of East Asian goats.

The domestic goat is one of the most important livestock species, but its origins and genetic diversity still remain uncertain. Multiple highly divergent maternal lineages of goat have been reported in previous studies. Although one of the mitochondrial DNA lineages, lineage B, was detected only in eastern and southern Asia, the geographic distribution of these lineages was previously unclear. Here, we examine the genetic diversity and phylogeographic structure of Asian goats by mitochondrial DNA sequences and morphological characteristics. The analyses of a total of 1661 Asian goats from 12 countries revealed a high frequency of lineage B in Southeast Asia. The frequency of this lineage tended to be higher in mountain areas than in plain areas in Southeast Asian countries, and there was a significant correlation between its frequency and morphological traits. The results suggest an original predominance of lineage B in Southeast Asia and the recent infiltration of lineage A into Southeast Asian goats.

Diversity and phylogeny of mitochondrial DNA isolated from mithun Bos frontalis located in Bhutan.

We sequenced the 16S rRNA gene in mitochondrial DNA to characterize mithun located in Bhutan and to increase our understanding of its origin. We compared mithun with yak, European cattle, Bhutanese zebu and Indian zebu. Sequencing revealed low nucleotide diversity within the mithun population and their phylogenetic proximity to gaur. A close relationship between Bhutanese mithun and gaur was confirmed by an additional comparison with wild gaur specimens from three locations in Bhutan. Direct domestication of mithun from gaur was supported, while maternal contribution from the cattle lineage during domestication was not supported.

Development of DNA markers for discrimination between domestic and imported beef.

In the meat industry, correct breed information in food labeling is required to assure meat quality. Genetic markers provide corroborating evidence to identify breed. This paper describes the development of DNA markers to discriminate between Japanese and Australian beef. Two Bos indicus-specific markers and MC1R marker were used as possible candidate markers. Amplified fragment length polymorphism method was employed to develop additional candidate markers. The 1564 primer combinations provided three markers that were converted into single nucleotide polymorphisms markers for high-throughput genotyping. In these markers, the allele frequencies in cattle from both countries were investigated for discrimination ability using PCR-RFLP. The probability of identifying Australian beef was 0.933 and probability of misjudgment was 0.017 using six selected markers. These markers could be useful for discriminating between Japanese and Australian beef and would contribute to the prevention of falsified breed labeling of meat.

Mitochondrial DNA variation and evolution of Japanese black cattle (Bos taurus).

This article describes complete mitochondrial DNA displacement loop sequences from 32 Japanese Black cattle and the analysis of these data in conjunction with previously published sequences from African, European, and Indian subjects. The origins of North East Asian domesticated cattle are unclear. The earliest domestic cattle in the region were Bos taurus and may have been domesticated from local wild cattle (aurochsen; B. primigenius), or perhaps had an origin in migrants from the early domestic center of the Near East. In phylogenetic analyses, taurine sequences form a dense tree with a center consisting of intermingled European and Japanese sequences with one group of Japanese and another of all African sequences, each forming distinct clusters at extremes of the phylogeny. This topology and calibrated levels of sequence divergence suggest that the clusters may represent three different strains of ancestral aurochs, adopted at geographically and temporally separate stages of the domestication process. Unlike Africa, half of Japanese cattle sequences are topologically intermingled with the European variants. This suggests an interchange of variants that may be ancient, perhaps a legacy of the first introduction of domesticates to East Asia.

Visual methods to assess cold fingers and experimental verification.

Cold fingers is complaint of many people. To independently assess actual finger temperature, this paper uses prototype sensors to capture blood vessel width and blood flow rates. We verify their feasibility for future home healthcare use along with far infrared camera outputs. We elucidate the impact of three remedies, massage, hot cocoa, and shoulder exercises, on 7 subjects.

cDNA cloning of pig testicular lactate dehydrogenase-C, thermal stability of the expressed enzyme, and polymorphism among strains.

Pig testicular lactate dehydrogenase-C (LDHC) cDNA was cloned and sequenced. The deduced sequence of 332 amino acids from pig LDHC shows 73% and 67% identity with that of pig LDHA (muscle) and LDHB (heart) respectively, whereas pig LDHA and LDHB isozymes shows 74% sequence identity. Pig and mouse LDHC cDNAs were subcloned into bacterial expression vector, and the expressed pig LDHC isozyme was shown to be as thermally stable as mouse LDHC isozyme. Pig genomic DNAs from Chinese Meishan, English Yorkshire, Danish Landrace and American Duroc were shown to exhibit polymorphic sites for restriction enzymes EcoRI, BamHI and PstI.

Sequences of the lizard cDNAs encoding lactate dehydrogenase (LDH) isozymes A (muscle) and B (heart).

The nucleotide and deduced amino acid sequences of cDNAs encoding L-lactate dehydrogenase (LDH) isozymes A (muscle) and B (heart) from the lizard, Sceloporus undulatus, were determined. The evolutionary relationships among LDH isozymes from animals, plants and bacteria are presented.

Reconstruction of axonal trajectory of individual neurons in the spinal cord using Golgi-stained serial sections.

A procedure bringing the axonal and dendritic cut ends found in the confronting planes of the serial sections face to face by means of serial photomicrographs provides a means for precisely reconstructing individual Golgiimpregnated neurons. Utilizing this method, it was possible to follow at the caudal level of a kitten's spinal cord the total course of the axons and dendrites of sixteen intramedullary neurons. The neurons located in the dorsal horn (laminae IV and V) send their long ascending axon into the lateral funiculus of the ipsi- or contralateral side, and, in one case, into the anterior funiculus of the ipsilateral side, giving off on the way several collaterals to the dorsal half of lamina VII of the ipsi- or contralateral side. Some of the neurons found in the intermediate region and the ventral horn (laminae VII, VIII and X) distribute their axonal branches in the intermediate region or the ventral horn of the ipsi- or contralateral side or both sides, running throughout their course in the grey matter, while the others act in the same manner by way of the lateral or anterior funiculi. Axonal and collateral endings terminate with small terminal knobs. The total axonal length of the neurons observed, with the exception of some with a long ascending trajectory, varies from 1,680 mum to 6,480 mum.

Morphological analysis of astrocytes in the bullfrog (Rana catesbeiana) spinal cord with special reference to the site of attachment of their processes.

An attempt was made to elucidate the morphological features of astrocytes in the bullfrog spinal cord by means of a combination of electron microscopy, the Golgi method, and the intravascular dye injection method. Astrocytic somata are densely concentrated both in the wall of the central canal and in its proximity, and diffusely distributed in the intermediate and the ventral part of the gray matter. The most complicated and densest vascular network is found in the dorsal part of the gray matter. There is little correlation between the density of the distribution of astrocytic somata and that of the vascular network. Each astrocyte emits one process and ramifies by repeated bifurcations as it approaches the white matter or enters it. All these branching processes reach the pial surface of the spinal cord (the principal processes). Total rostrocaudal extent of their ramification is within 400 micron. A great number of small lateral offshoots (the secondary processes) arise both from the somata and the principal processes. Electron microscopy of vessel walls and the pial surface revealed that the principal processes attach to the subpial basement membrane with a specialized structure, an electron-dense layer, while the secondary processes merely surround blood vessels in a mode of juxtaposition. Comparison between amphibian and mammalian astrocytes is made regarding the site of attachment of their processes.

Reconstruction of neurons of dorsal horn proper using Golgi-stained serial sections.

Three neurons located in the dorsal horn (Lamina III) have been reconstructed using Golgi-stained serial sections. These cells including dendritic and axonal branchings appear to remain within the dorsal horn and to belong to small-sized local interneurons of the dorsal horn proper. The measurements have been made with various criteria: the medio-lateral, dorso-ventral and rostro-caudal extents of the dendritic tree as well as the axonal branchings, total dendritic and axonal length, number of axonal endings and distance from the starting point of the axon at the dendrite to the point where the dendrite joins the soma (S-A distance).

Reconstruction of trajectory of primary afferent collaterals in the dorsal horn of the cat spinal cord, using Golgi-stained serial sections.

Using Golgi-stained serial sections obtained at the sacro-caudal levels of the cat spinal cord, it was possible to reconstruct the trajectory of primary afferents. They were classified into two groups: reliable primary afferents directly traced from the dorsal root and probable primary afferents traced from the dorsal funiculus or Lissauer's tract. The diameters of the reliable primary afferents vary from 0.88-1.88 mum. According to their courses, reliable primary afferents as well as probable primary afferents were classified into three groups: the first is distributed to both medial and lateral halves of the dorsal horn, the second to the medial half, and the third to the lateral half. Commissural fibers were also observed among the probable primary afferents. The rostro-caudal and medio-lateral extents of reliable primary afferents are found to be between 250 and 950 mum and 270 and 700 mum respectively, while those of the probable primary afferents were between 125 and 670 mum and 270 and 1,640 mum respectively. These primary afferent fibers are connected with at least two or more laminae of the dorsal horn gray matter.

Trajectory of group Ia afferent fibers stained with horseradish peroxidase in the lumbosacral spinal cord of the cat: three dimensional reconstructions from serial sections.

A reconstruction was made of the intramedullary trajectory of 23 physiologically identified Ia afferents from cat hind limb muscles (medial gastrocnemius, soleus, plantaris, flexor digitorum-hallucis longus, and hamstring). The afferents were stained by intra-axonally injected HRP. The axons of these afferents were traced over distances of 5.8 mm to 15.7 mm rostrocaudally. In the dorsal funiculus fibers from all the muscles showed a similar course and similarly bifurcated into an ascending and a descending branch. The mean diameters of stem axons, ascending branches, and descending branches were 6.6 micrometer, 5.8 micrometer, and 3.0 micrometer, respectively. Within the analyzed lengths of the spinal cord five to eleven collaterals were given off from the two branches. The distances between adjacent collaterals of the ascending and descending branches averaged 1200 micrometer and 790 micrometer, respectively. The collaterals as a rule passed through the medial half of the dorsal horn before they entered the deeper parts of the gray matter. The terminal distribution areas common to all Ia collaterals were: (1) the medial half of the base of the dorsal horn, mainly lamina VI: (2) lamina VII; and (3) lamina IX. The numbers of terminals were largest in lamina IX and smallest in lamina VII. The density of terminals in lamina IX was highest in the homonymous motor cell column. The terminal distribution areas of adjacent collaterals showed no overlap in the sagittal plane. Terminal branches carried one bouton terminal and up to six boutons en passage with an average of 1.8 terminals per terminal branch. Apparent axosomatic and axodendritic contacts were seen on small-sized and medium-sized neurons in laminae V-VI, medium-sized neurons in lamina VII, and large neurons in lamina IX. One motoneurons was contacted by an average of 3.3 terminals. In addition to the common features, Ia collaterals of various muscles of origin showed some differences in their trajectories in the ventral horn, and in their terminations in the gray matter.

Morphology of expiratory neurons of the Bötzinger complex: an HRP study in the cat.

In anesthetized and artificially ventilated cats, the physiological and morphological properties of expiratory neurons or their axons of the Bötzinger complex (BOT) were studied using intracellular recording and intracellular HRP labeling techniques. Thirteen expiratory neurons (nine cell somata and four axons) were successfully stained. Four of them were motoneurons, having relatively large cell somata in the retrofacial nucleus (RFN) and axons without any collaterals inside the brainstem. All the motoneurons showed a plateau shape of depolarization potentials during the expiratory phase. Any of the other nine expiratory neurons exhibited augmenting type firing or membrane potential changes during the expiratory phase. In five out of nine augmenting neurons, cell somata were stained and located ventral to the RFN. In four, only axons were stained. The majority of the augmenting neurons had two major axonal branches: one traveling toward the contralateral side and the other descending ipsilaterally in the brainstem. The most striking feature of the axonal trajectory was that all of the stained augmenting expiratory neurons, including the axons, had collateral branches with synaptic boutons in the BOT area, thus indicating that BOT expiratory neurons interact with some respiratory neurons in the BOT area and its vicinity.

Morphology of single primary vestibular afferents originating from the horizontal semicircular canal in the cat.

The central projections of physiologically characterized vestibular nerve fibers originating from the horizontal semicircular canal were studied in the vestibular nuclei of adult cats after intracellular staining with horseradish peroxidase (HRP). First, primary nerve fibers were physiologically classified as regular or irregular types on the basis of the regularity of the spontaneous discharge pattern. Then, these two types of fibers were morphologically analyzed and compared following HRP intraaxonal injection. The two types of axons showed a basically similar trajectory in the four major vestibular nuclei. They bifurcated into an ascending and a descending branch in the ventrolateral part of the lateral vestibular nucleus (LVN). The ascending branch extended rostrally and gave off one or two collaterals in the superior vestibular nucleus (SVN), although some of the ascending branches further ran rostrally into the cerebellum. The collaterals, while running medially, gave rise to fine terminal branches with en passant boutons in the SVN, and further coursing caudally, they entered the rostral part of the medial vestibular nucleus (MVN). The descending branch, while running caudally in the lateral part of the LVN and the inferior vestibular nucleus (IVN), gave off several thick collaterals to the MVN and extensive terminals were present in the IVN and MVN. In each primary axon, about one-third of the total terminal boutons were distributed in each of the SVN, the MVN, and the IVN. In contrast to this similarity of the overall axonal trajectory within the vestibular nuclei, both types of axons exhibited several marked differences in diameter and in the mode of terminal arborization. In almost every part of the ramification, the irregular-type fibers were thicker than the regular-type fibers. In the regular-type axons, many small terminal boutons (mean size, 2.4 x 1.4 microns, N = 2,739) were located in close proximity (100-150 microns) to the parent collateral. In the irregular-type axons, slightly larger terminal boutons (mean size, 3.0 x 1.7 microns, N = 1,287), were spread more widely (200-300 microns) around their collaterals. These clear morphological differences between the regular-type and the irregular-type terminal axons were consistently observed in any vestibular nucleus.

Morphology of augmenting inspiratory neurons of the ventral respiratory group in the cat.

The present study examined, in Nembutal-anesthetized and artificially ventilated cats, the morphologic properties of the inspiratory neurons of the ventral respiratory group (VRG). Horseradish peroxidase (HRP) was injected into 21 augmenting inspiratory or late inspiratory neurons with peak firing rates in the late inspiratory phase. The majority of the stained neurons were antidromically activated by stimulation of the cervical cord. Thirteen somata, located within or around the nucleus ambiguus (AMB), between 100 microns caudally and 2,000 microns rostrally to the obex, were stained. In ten cases, the stem axons issuing from the cells of origin coursed medially to cross the midline without giving off any axonal collaterals. Three neurons gave rise to axonal collaterals on the ipsilateral side, distributing boutons in the medullary reticular formation, in the vicinity of the AMB, hypoglossal nucleus, solitary tract, and dorsal motor nucleus of the vagus. In eight neurons, only the axons were labeled; in four of these, which were antidromically activated from the spinal cord, the stem axons crossed the midline 2,000-3,000 microns rostral to the obex and descended in the reticular formation around the AMB down to the cervical cord. They issued several axonal collaterals, distributing terminal boutons at the level of the caudal end of the retrofacial nucleus and about 1,000 microns rostral and caudal from the obex. Terminals were found mainly in and around the AMB, and a few were found in the vicinity of the dorsal motor nucleus of the vagus. The remaining four nonactivated axons distributed their terminal boutons widely in the reticular formation around the AMB. Thus, the augmenting inspiratory neurons of the VRG were shown to project not only to the spinal cord, but also to the VRG, hypoglossal nucleus, and dorsal motor nucleus of the vagus.

Trajectory of group Ia and Ib fibers from the hind-limb muscles at the L3 and L4 segments of the spinal cord of the cat.

Nineteen physiologically identified group Ia and five group Ib fibers at the L3 and L4 levels of the spinal cord originating from various hind-limb muscles were intraaxonally injected with horseradish peroxidase (HRP). The trajectories of the stained axons were reconstructed. They extended for distances of 8.6 mm-18.0 mm rostrocaudally. Ascending axons ran in various regions of the dorsal funiculus: The ascending axon from a toe muscle (3 microns in diameter) ran in the ventral most part of the paramedian region; those from shank muscles (3.0-5.0 microns) in both dorsal and ventral paramedian regions; those from thigh muscles (5.0-7.0 microns) in both the paramedian and the more lateral regions; and those from hip muscles (6.0-7.0 microns) in the lateral region. Main collaterals arising from the parent fiber were given off at intervals of 0.5-6.2 mm (mean 2.4 mm). Collaterals of a fiber from a toe muscle (1.0 micron in diameter) entered Clarke's column from the dorsomedial side and ramified mostly in the dorsomedial one-third of the column. Collaterals of fibers from shank muscles (1.0-2.0 microns) entered Clarke's column from the dorsal side and terminated in its middle parts as well as in laminae V-VII. Collaterals of fibers from thigh muscles (1.0-2.5 microns) passed lateral to or through the lateral part of Clarke's column and terminated in its ventrolateral part and in laminae V-VIII. Collaterals of fibers from hip muscles (1.5-2.5 microns) passed lateral to Clarke's column and ramified mostly in laminae VII-IX. As the muscle of origin became more proximal, the proportion of termination outside of Clarke's column progressively increased. Thus, the trajectory of group I fibers was somatotopically organized both in the dorsal funiculus and in the gray matter. The long axis of boutons ranged from 0.5 to 17 microns in Ia fibers and from 0.5 to 8 microns in Ib fibers. "Giant" Ia boutons (above 7 microns) were found both in and outside Clarke's column.

Chicken ornithine transcarbamylase gene, structure, regulation, and chromosomal assignment: repetitive sequence motif in intron 3 regulates this enzyme activity.

Ornithine transcarbamylase (OTC) is one of the urea cycle enzymes. While the chicken is a uricotelic animal, it has measurable OTC activity in its kidney. OTC activity is highly variable within and between chicken breeds. Chicken OTC may have some physiological significance because of its significant activity in the kidney. We cloned the OTC cDNA from chicken kidney and found 77% homology between the deduced amino acid sequence of the mature protein and that of mammals. The chicken OTC gene spans 26 kb, consists of 10 exons and 9 introns, and utilizes the same exon-intron boundaries as the human gene. The 5'-flanking region contains a putative TATA box and two potential regulatory sites, but neither the 5'-flanking region nor the splice sites correlated with variation in OTC activity. In intron 3, two polymorphic sites were found: one comprising a deletion of 401 nucleotides; and the other was a length and sequence polymorphic region located 8 bases upstream from the deletion. The latter polymorphism provides an explanation for phenotypic variation in OTC. Linkage analysis has suggested reassignment of the chicken OTC gene from the suggested Z chromosome to chromosome 1q.

Morphological analysis of single astrocytes of the adult cat central nervous system visualized by HRP microinjection.

The morphology of individual astrocytes of the adult cat was analyzed by HRP microinjection and light microscopy. The astrocytes had generally two types of processes: (1) thread-like processes of relatively constant width with few ramifications and few lamellar appendages and (2) the sinuous processes with clusters of lamellar appendages. The former processes were morphologically characterized as follows: (1) Those of fibrous astrocytes were frequently remarkably long, ranging from 600 to 1500 microns. They were much longer than any astrocytic processes hitherto reported in the literature. In contrast, those of protoplasmic astrocytes were usually short (30-400 microns), and were sometimes decorated with lamellae. (2) The processes often terminated in endfeet on the subpial surface of the brain tissue or on blood vessel walls. The number of endfeet per cell varied from 1-11 with a tendency to split into two populations close to each extreme number. Another type of endfoot was also found, i.e. swellings along the trunk of the processes which made side-to-side contact with the vessel wall. The sinuous processes rich in lamellae were predominant in protoplasmic astrocytes, and clearly corresponded to the sheet- or veil-like processes of Golgi-impregnated astrocytes. They formed an ellipsoidal field (100 microns for the longer, and 50-60 microns for the shorter, diameter) around the nucleus.

Electron microscopic comparison of the terminals of two electrophysiologically distinct types of primary vestibular afferent fibers in the cat.

Two types of electrophysiologically distinct vestibular primary afferents (regular-type and irregular-type) were injected intraaxonally with HRP and their terminals were compared using the electron microscope. In general, regular-type terminals were small and two or more terminals contacted a single dendrite side by side, whereas irregular-type terminals were large and covered a wide surface of the dendrites. Both types of terminals contained spherical clear vesicles with almost the same diameter. Prominent postsynaptic membrane specializations were observed in both types. Mitochondria in both types had an elliptical profile with the same elongation index, but mitochondria in the regular-type terminals were thicker than those in the irregular-type ones.