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R Elde

Publications and source records attributed to R Elde.

At least 73 records · Page 4Linked to original sources

Organization of the serotonergic innervation of spinal neurons in rats--III. Differential serotonergic innervation of somatic and parasympathetic preganglionic motoneurons as determined by patterns of co-existing peptides.

The spinal cord is innervated by brainstem serotonergic neurons, some of which contain substance P and/or thyrotropin-releasing hormone in addition to serotonin. These neurons project at least three types of axons to the spinal cord: those containing both substance P and thyrotropin-releasing hormone, those containing thyrotropin-releasing hormone but not substance P, and those containing neither substance P nor thyrotropin-releasing hormone. However, the organization of the different types of serotonergic processes is unclear. In the present studies, the types of serotonergic axons projecting to two kinds of spinal neurons were examined. Somatic and parasympathetic preganglionic motoneurons were labeled retrogradely from the pelvic or sciatic nerve, respectively. Sections containing these neurons were stained either for serotonin and substance P, or for serotonin and thyrotropin-releasing hormone. Of a total of 428 profiles examined that were retrogradely labeled from the sciatic nerve, 425 (99%) were apposed by serotonin-immunoreactive varicosities; similarly, of a total of 382 profiles examined that were retrogradely labeled from pelvic nerve, 353 (92%) were apposed by serotonin-immunoreactive varicosities. However, differences appeared to exist between the types of serotonergic varicosities innervating these two groups of neurons. Among the profiles labeled from the sciatic nerve, it was estimated that over 97% were apposed by serotonin-immunoreactive varicosities in which serotonin co-existed with substance P and thyrotropin-releasing hormone. In contrast, among the profiles labeled from pelvic nerve that were apposed by serotonin-immunoreactive varicosities, it was estimated that less than 1% were apposed by serotonin-immunoreactive varicosities containing both thyrotropin-releasing hormone and substance P. We estimate that most of the remainder (about 80%) were apposed by serotonin-immunoreactive varicosities containing thyrotropin-releasing hormone but not substance P. We conclude that both the cell bodies of neurons retrogradely labeled from the pelvic nerve and those labeled from the sciatic nerve were apposed by serotonin varicosities. However, these two systems of neurons appear to be innervated largely by two different populations of serotonergic cells. This suggests that the raphe-spinal serotonergic system may independently modulate the activities of somatic motoneurons and parasympathetic preganglionic motoneurons.

Animals↗

Regional distribution of serotonin and substance P co-existing in nerve fibers and terminals in the brainstem of the rat.

Two-color fluorescence immunohistochemistry was used to identify and map the distribution of nerve processes immunoreactive for both serotonin and substance P in the rat brainstem. Doubly labeled fibers were observed throughout the brainstem, but tended to be densest in cranial nerve motor nuclei and in reticular regions of the ventral medulla. In the trigeminal motor nucleus, the facial nucleus and the spinal accessory nucleus, the majority of serotonergic varicosities also appeared to contain substance P; in the occulomotor nucleus and the hypoglossal nucleus the numbers of double-labeled and single-labeled serotonergic varicosities were roughly equal. Thus, co-existence of substance P with serotonin was common in many cell groups innervating skeletal muscle. The proportion of double-labeled varicosities was significantly lower in the nucleus of the solitary tract, wherein single-labeled varicosities were much more common. Double-labeled fibers and varicosities were also significantly less common in the spinal trigeminal nucleus. In addition, double-labeling appeared to be uncommon in regions involved in the processing of special sensory information (e.g. auditory, vestibular and visual pathways). These results demonstrate a subpopulation of serotonergic fibers immunoreactive for substance P in the brainstem of the rat. The consistently high density of double-labeled processes in cranial nerve motor nuclei suggests that, as may be the case in the spinal cord, neurons containing serotonin and substance P regulate the activity of motoneurons that innervate skeletal muscle. In addition, they may be involved in other aspects of the function of the brainstem.

Animals↗

Dopamine transporter mRNA in neurons of the rat hypothalamus.

The membrane-bound dopamine transporter (DAT) protein terminates dopaminergic neurotransmission by rapid reaccumulation of dopamine into presynaptic nerve endings. The distribution of neurons expressing DAT mRNA was investigated in the rat hypothalamus using in situ hybridization histochemistry and oligonucleotide probes to sequences of rat DAT mRNA. DAT mRNA-containing cell bodies were demonstrated in the ventral aspect of the periventricular nucleus (A14 dopamine cell group), in the zona incerta (A13), in the dorsomedial part of the arcuate nucleus (A12), and in scattered regions of the posterior hypothalamus. There was no labelling in magnocellular neurons of the supraoptic or paraventricular nuclei, or in neurons of the ventrolateral division of the arcuate nucleus, areas previously shown to contain the dopamine-synthesizing enzyme tyrosine hydroxylase. The results suggest that hypothalamic dopamine neurons have an active uptake mechanism for dopamine, however, that DAT mRNA levels may be considerably lower than those encountered in the ventral midbrain.

Animals↗

Regulation of an opioid-binding protein in NG108-15 cells parallels regulation of delta-opioid receptors.

An opioid-binding protein has recently been purified from bovine brain and cloned, and its cDNA sequence has been obtained. Indirect evidence suggests that this protein has a role in opioid-receptor function. However, because direct testing of its function by expression of its cDNA has not yet been possible and because its structure bears no resemblance to G protein-coupled receptors, the role of this protein in opioid-receptor activity is still in question. An antibody raised to a portion of the predicted amino acid sequence of opioid-binding cell-adhesion molecule (OBCAM) specifically labeled the surface of NG108-15 cells, as visualized by immunofluorescence with confocal microscopy. Furthermore, chronic treatment of these cells with opioid agonist, which down-regulates opioid receptors, reduced OBCAM immunoreactivity (ir). Down-regulation of both opioid receptors and OBCAM-ir was greatest after chronic treatment of NG108-15 cells with delta-opioid agonists, as well as with nonselective agonists such as etorphine, whereas other agonists including [D-Ala2-N-MePhe4-Gly-ol]enkephalin, morphine, levorphanol, dynorphin A-(1-13), and U-50,488H were less effective or ineffective. Chronic treatment of NG108-15 cells with muscarinic agonists had no effect on OBCAM-ir. Furthermore, NG108-15 cells transfected with an antisense construct to OBCAM have a reduced density of opioid-binding sites as well as reduced OBCAM-ir. Taken together, these results strongly suggest that OBCAM has a role in opioid-receptor function in NG108-15 cells.

Amino Acid Sequence↗

Identification of neurons expressing thyrotropin releasing-hormone receptor mRNA in spinal cord and lower brainstem of rat.

The distribution of mRNA coding for a pituitary thyrotropin releasing-hormone (TRH) receptor was examined on sections of spinal cord and lower brainstem of rat using in situ hybridization. Hybridization signals were observed over large neurons in the ventral horn in cervical, thoracic, and lumbar segments of spinal cord, and over neurons in the motor nuclei of the lower brainstem. Although significant thyrotropin-releasing hormone binding has been reported in the superficial dorsal horn, only background levels of hybridization were observed over neurons in this region. These findings suggest that mRNA coding for thyrotropin-releasing hormone receptor is expressed in some spinal and brainstem motor neurons. Since many of these neurons are innervated by TRH-containing afferents, TRH may exert a direct effect upon at least some of these cells.

Animals↗

Distribution of thyrotropin-releasing hormone receptor messenger RNA in the rat brain: an in situ hybridization study.

Based on the recent cloning of the mouse thyrotropin-releasing hormone receptor, oligonucleotide probes complementary to the DNA sequence were constructed and used for in situ hybridization studies on the rat brain. Thyrotropin-releasing hormone receptor messenger RNA was found in many areas of the brain, mostly showing high degree of overlap with the distribution thyrotropin-releasing hormone binding sites as previously revealed in autoradiographic studies. Thus, a strong signal was observed in the accessory olfactory bulb, the perirhinal sulcus, the ventral aspects of the hippocampal formation, some amygdaloid nuclei, the diagonal band nucleus, parts of nucleus accumbens, the bed nucleus of the stria terminalis, dorsomedial, lateral and perifornical hypothalamic regions, the septohippocampal nucleus, parts of the vestibular complex, as well as many bulbar motoneurons including the facial, dorsal vagal, ambiguus and hypoglossal nuclei, the superficial layer of the spinal trigeminal nucleus, and motoneurons and dorsal horn neurons in the spinal cord. Cells within one and the same nucleus expressed varying levels of thyrotropin releasing hormone receptor messenger RNA suggesting marked differences in rate of receptor synthesis. Most of these areas receive an input by thyrotropin-releasing hormone-positive nerve endings. Taken together these results suggest that thyrotropin-releasing hormone receptors are mostly localized in the vicinity of the cell bodies which express thyrotropin-releasing hormone receptor messenger RNA and mediate the wide range of actions that have been recorded after administration of exogenous thyrotropin-releasing hormone.

Animals↗

Expression of acidic and basic fibroblast growth factors in the substantia nigra of rat, monkey, and human.

The distribution of acidic (aFGF) and basic (bFGF) fibroblast growth factor mRNA and protein were examined in mesencephalon by immunohistochemistry, immunoblot analysis, in situ hybridization histochemistry, and RNA analysis. Coexistence of aFGF or bFGF with tyrosine hydroxylase protein in nigral cells was observed with immunohistochemistry. Both aFGF and bFGF mRNAs were found in the substantia nigra. Unilateral 6-hydroxydopamine lesions of nigrostriatal neurons resulted in a loss of aFGF and tyrosine hydroxylase [L-tyrosine, tetrahydropteridine: oxygen oxidoreductase (3-hydroxylating), EC 1.14.16.2] mRNA-positive neurons on the lesioned side. The distribution of aFGF mRNA in monkey brain was similar to that seen in the rat. RNA and immunoblot analysis confirmed the presence of both aFGF and bFGF mRNAs and proteins in the substantia nigra of rat, monkey, and human.

Animals↗

Detection and characterization of a sensory microganglion associated with the spinal accessory nerve: a scanning laser confocal microscopic study of the neurons and their processes.

The spinal accessory nerve has been generally thought to be a cranial nerve with purely motor function, innervating the trapezius and sternocleidomastoid muscles. The present study identified clusters of sensory neurons consistently associated with this cranial nerve in adult rats. Either a single microganglion or several dispersed microganglia were found that adhered to the spinal root of the nerve, to small vessels, or were free within the subarachnoid space. The neurons of the ganglion had axons that joined the spinal root of the nerve proximal to its exit from the skull. Additional branches appeared to have an intracranial distribution within the arachnoid of the brainstem and along its vessels. Several findings suggest that the function of the ganglion is sensory and not autonomic. First, the architectural features of neurons within the ganglion (including their size, pseudounipolar morphology, and the lack of synaptic contacts) are similar to those of neurons in other sensory ganglia. Second, substance P and calcitonin gene-related peptide coexist within neurons of the microganglion, whereas markers for the major transmitters found in autonomic ganglia in rats are absent. Third, the expression of peptides in neurons of the ganglion was sensitive to neonatal capsaicin treatment. Finally, neurons within the ganglion were filled with a retrogradely transported dye after injection of the dye into the cervical spinal cord. Although the function of the ganglion is not known, its features are consistent with a role in nociception from the muscles of the spinal accessory complex, and it may be involved in headaches that have an occipital distribution.

Accessory Nerve↗

Distribution of CGRP-, VIP-, D beta H-, SP-, and NPY-immunoreactive nerves in the periosteum of the rat.

In light of the possible role peripheral nerves may play in bone metabolism, the morphology of calcitonin gene-related peptide (CGRP)-, vasoactive intestinal peptide (VIP)-, substance P (SP)-, neuropeptide Y (NPY)-, and dopamine-beta-hydroxylase (D beta H)-immunoreactive nerve fibers was examined in whole-mount preparations of periosteum of membranous bones (calvaria, mandible) and long bones (tibia) from the rat. Periosteum from animals treated to remove selectively either the sympathetic or fine-caliber primary afferent nerves was also examined to determine the origin of the nerve fibers. We found a consistent and often dense innervation of the periosteum. The innervation patterns of the calvaria and mandible were similar, with networks of nerves spread across the surface of the bone. Nerves in the tibial periosteum were oriented in the longitudinal axis and were more numerous at the epiphyses than in the mid-shaft region. CGRP-immunoreactive fibers were widely and densely distributed. The presence of populations of CGRP-immunoreactive fibers of differing calibers and perivascular arrangements suggests that such nerves in bone tissues may serve different functions. SP-immunoreactivity was present in a fine network of varicose fibers in the superficial layers of the periosteum. CGRP- and SP-immunoreactive nerve fibers were dramatically reduced in periosteum of capsaicin-treated animals as compared to controls, indicating the sensory origin of these nerves. VIP-immunoreactive nerve fibers were distributed in the periosteum of mandible and calvaria as small networks and individual fine varicose fibers. In tibial periosteum, larger networks of these fibers were visible. VIP-immunoreactive nerve fibers in the periosteum were associated with both vascular and nonvascular elements within the layers of cells closest to the bone, suggesting that VIP may serve more than one function in periosteal tissues. NPY-immunoreactive fibers were largely confined to vascular elements; occasional fibers were observed among the bone-lining cells. D beta H-immunoreactivity was associated only with blood vessels. VIP-, NPY-, and D beta H-immunoreactivities were dramatically reduced in the periosteum of guanethidine-treated animals, indicating the sympathetic origin of these nerves.

Animals↗

Effects of neonatal sympathectomy and capsaicin treatment on bone remodeling in rats.

Bone metabolism may be influenced by the innervation of skeletal tissues. Neuropeptides such as vasoactive intestinal peptide, from sympathetic nerves, and calcitonin gene-related peptide, from sensory nerves, have been implicated as local modulators of bone metabolism. The effect of neonatal sympathectomy and of capsaicin-induced sensory denervation in rats was studied on the following: (i) the radial bone growth and apposition rate in tibiae (normal growth and modeling) and (ii) the percentage of periosteal surface of the mandible occupied by osteoclasts during induced remodeling. Neonate rats were treated with guanethidine, capsaicin, or appropriate vehicle. At seven weeks, maxillary molars were removed to induce remodeling on the buccal surface of the mandible. Animals were killed four days after surgery. Cross-sectional cortical area, medullary area, and periosteal apposition rate were measured by histomorphometry in ground sections of tibiae. The percentage of periosteal surface at the remodeling site occupied by osteoclasts (stained for acid phosphatase) was measured in frozen, undecalcified sections. There was no significant difference in cortical or medullary area or periosteal apposition rate in tibiae between each drug treatment and its control. However, the mandibular bone surface occupied by osteoclasts was increased 45.5% (P less than or equal to 0.005) in animals treated neonatally with guanethidine compared to controls. In contrast, the mandibular surface occupied by osteoclasts was decreased 21.2% (P less than or equal to 0.04) in animals treated neonatally with capsaicin compared to controls. The alteration of bone remodeling (osteoclast surface) by both treatments indicates that sensory and sympathetic nerves play a role in focal metabolism of bone.

Animals↗

Prominent expression of acidic fibroblast growth factor in motor and sensory neurons.

Several growth factors originally characterized and named for their action on a variety of cells have more recently been suggested to be importantly involved in the development and maintenance of the nervous system. Acidic fibroblast growth factor (aFGF) is a member of a family of seven structurally related polypeptide growth factors. The cells responsible for expression of aFGF in the nervous system of adult rats have been identified using an affinity-purified antibody to aFGF in immunohistochemical studies and synthetic oligonucleotide probes for in situ hybridization studies. High levels of aFGF expression were observed in motoneurons, primary sensory neurons, and retinal ganglion neurons. Glial cells did not express detectable amounts of aFGF. Confocal and electron microscopic analysis suggested that a large portion of aFGF immunoreactivity was associated with the cytoplasmic face of neuronal membranes, consistent with the hypothesis that aFGF is a sequestered growth factor.

Animals↗

Localization of neuropeptide receptor mRNA in rat brain: initial observations using probes for neurotensin and substance P receptors.

The expression of receptors for neurotensin and substance P was examined in rat brain and spinal cord using in situ hybridization with synthetic oligonucleotide probes. Strong hybridization signals for neurotensin receptor mRNA were observed over neurons i.a. in the diagonal band, medial septal nucleus, nucleus basalis magnocellularis, suprachiasmatic nucleus, supramammillary area, substantia nigra and ventral tegmental area. Strong hybridization signals for substance P receptor mRNA were observed over scattered, large neurons in the striatum, and in the spinal cord over neurons in the dorsal horn, the area around the central canal and preganglionic autonomic neurons. Thus, discrete neurons in several brain regions express a G-protein-coupled receptor with which endogenous neurotensin and substance P may interact.

Animals↗

An improved method for preparing cryostat sections of undecalcified bone for multiple uses.

We developed a technique that permits the use of serial sections (7-20 microns) from a single fixed piece of bone tissue for immunofluorescence, measurement of fluorescent bone labels, enzyme histochemistry, and general staining. This technique combines modifications of previously established methods with perfusion of the polymer polyvinylpyrrollidone (PVP) to improve sectioning, and produces reliable sections with good preservation of both hard and soft tissues. The combination of techniques from several workers, the use of perfusion with a polymer to increase the sectionability of the bone, and the addition of a gelatin adhesive on top of pressure-sensitive adhesives represent a significant improvement over previously described methods. The sections obtained are usable for immunocytochemistry, general staining, enzyme histochemistry, and visualization of fluorescent bone labels. We have consistently used tissues prepared in this manner for immunohistochemical demonstration of neuropeptides in skeletal tissues and for localizing tartrate-resistant acid phosphatase (TRAP). In addition, other tissues obtained from PVP-perfused rats, such as brain, spinal cord, muscle, gut, and sympathetic ganglia, are also well preserved and demonstrate immunohistochemical staining comparable to and possibly superior to that obtained with normal fixation protocols.

Animals↗

Vasoactive intestinal peptide distribution and colocalization with dopamine-beta-hydroxylase in sympathetic chain ganglia of pig.

The extent to which vasoactive intestinal peptide (VIP) occurs in postganglionic cells of the sympathetic chain and whether these cells are noradrenergic, cholinergic, or neither is unknown. We have (1) established the extent of VIP-containing cell bodies and fibers in all levels of the sympathetic chain in pig and (2) determined the coexistence of VIP and dopamine-beta-hydroxylase (DBH) using immunofluorescence. Weanling pigs were perfusion fixed. Alternate serial frozen sections of cervical, thoracic and lumbar paravertebral ganglia were processed for immunofluorescence with antisera to VIP or DBH. VIP-immunoreactive cells were most evident in thoracic ganglia with a few occurring in stellate and lumbar ganglia. While the cells were distributed throughout individual thoracic ganglia, intensely stained cells consistently occurred just proximal to exiting fiber bundles and sent long processes into these bundles. VIP-immunoreactive fibers occurred in all ganglia. Fiber density varied at each level. Varicose fibers with VIP staining were found to be especially evident from the caudal cervicothoracic ganglion through upper lumbar ganglia, in many cases forming networks of beaded varicosities around principal ganglion cells. Fibers and cells were associated with both interganglionic connectives and rami. A majority of neurons in all ganglia were DBH-immunoreactive but with varying intensities. VIP- and DBH-immunoreactivity coexisted in the same perikarya in some cells in thoracic ganglia, preferentially in cells moderately DBH-immunoreactive. We conclude that VIP-immunoreactive cells are distributed primarily in the thoracic sympathetic chain ganglia and that fibers containing VIP are found at all ganglia levels. Additionally, some VIP-immunoreactive postganglionic neurons in paravertebral ganglia may be noradrenergic.

Animals↗

Distribution of cholecystokinin-immunoreactive cell bodies in the male and female rat: II. Bed nucleus of the stria terminalis and amygdala.

The distribution of cholecystokinin-immunoreactive (CCK-I) cell bodies was studied in the bed nucleus of the stria terminalis (BST) and amygdaloid complex of colchicine-treated male and female rats. Immunoreactive cells were visualized in the BST medial amygdaloid (MeA), central lateral, basolateral, basolateral ventral, medial, intercalated, anterior cortical, and posterior cortical nuclei and the amygdalohippocampal zone. Several significant sex differences were observed. In the male, a dense aggregation of CCK-I cell bodies was visualized in the MeA, especially in the dorsocaudal part and in the encapsulated part of the BST. In comparison, female rats had relatively fewer immunoreactive cells in both of these regions. In the lateral and basolateral amygdaloid nuclei, however, more CCK-I cells were visualized in the female than in the male, but the difference was not statistically significant. These data provide characterization of a sexually differentiated CCK system. In addition, we observed that the number of CCK-I cells in the BST and posterodorsal part of the MeA was substantially reduced after castration. The number of CCK-I cells in female rats, however, was not significantly reduced after ovariectomy in any of the regions studied. These findings imply that the steroid regulation of CCK is sexually differentiated. The sexually dimorphic distribution of CCK-I cells in areas that are targets of steroid hormones and regulate reproductive processes is consistent with the possibility that CCK participates in central integration of sensory and steroidal input that modulates reproductive behavior.

Amygdala↗

Calcitonin gene-related peptide-immunoreactive nerve fibers in mandibular periosteum of rat: evidence for primary afferent origin.

Peptidergic neurons may play a role in the local regulation of bone mineralization. The neuropeptide vasoactive intestinal peptide (VIP) increases bone resorption in vitro, while calcitonin gene-related peptide (CGRP) has been shown to inhibit bone resorption in vitro. We have previously reported that sympathetic nerves with VIP-immunoreactivity innervate bone and periosteum. In the present study we sought to determine if CGRP fibers, like VIP fibers, exist in periosteum and what their origin might be. In whole-mount preparations of mandibular periosteum from rat, CGRP- and VIP-immunoreactive (IR) nerve fibers were present as networks within the periosteum. In preparations using two-color immunofluorescence, most CGRP-IR fibers were also immunoreactive for substance P (SP). In rats in which the subperiosteal space subjacent to the mandibular molars was injected with Fast blue or Fluoro-gold, retrogradely labeled cells were seen in ipsilateral trigeminal ganglia, superior cervical ganglia, and nodose ganglia. Individual cells labeled with both CGRP immunoreactivity and retrograde tracer were seen only in the mandibular portion of the trigeminal ganglion. These data suggest that CGRP-IR nerve fibers in periosteum may be of primary afferent origin. Given the reported effects of CGRP on bone mineralization, the present results suggest that primary afferent nerves containing CGRP and SP, as well as sympathetic nerves containing VIP, may play a role in focal bone remodeling.

Afferent Pathways↗

Expression of eight neuropeptides in intraocular spinal cord grafts: organotypical and disturbed patterns as evidenced by immunohistochemistry.

The present study examines the distribution of several neuropeptides, as revealed by immunohistochemistry in the isolated cord. Fetal rat spinal cord was grafted to the anterior chamber of the adult Sprague-Dawley albino rats. After intraocular maturation for 2-3 months, the amount and distribution of somatostatin, neuropeptide Y, substance P, enkephalin, vasoactive intestinal peptide, peptide histidine-isoleucine, calcitonin gene-related peptide and cholecystokinin immunoreactive terminals and cell bodies were analysed using indirect fluorescence immunohistochemistry. The visualization of immunoreactive cell bodies in the grafts was enhanced using a novel intraocular colchicine treatment. In the graft a rich network of somatostatin-positive terminals was found with a high density in well-demarcated areas reminiscent of substantia gelatinosa of the dorsal horn of normal spinal cord. A large number of small- to medium-sized somatostatin neurons was found throughout the grafts without colchicine treatment. This is in contrast to normal spinal cord, where positive neurons were difficult to visualize without colchicine and were mainly confined to the dorsal horn. Neuropeptide Y had a distribution in the grafts similar to that of somatostatin and neuropeptide Y cells were found throughout the grafts without colchicine treatment. In normal spinal cord, neuropeptide Y-positive fibers were found mainly in substantia gelatinosa with a sparse network in the ventral horn. Enkephalin-positive fibers were found throughout the grafts. The distribution of fibers resembled that of somatostatin and neuropeptide Y with distinct zones of high fiber density in well-demarcated areas, whereas the density of nerve fibers in the rest of the graft neuropil was moderate to low. The distribution of substance P was similar to that of enkephalin. After colchicine treatment, both enkephalin- and substance P-positive cell bodies were visualized. In the intact spinal cord both peptides were seen in the entire gray matter with the highest concentrations in the superficial laminae of the dorsal horn. Antisera against calcitonin gene related-peptide, revealed a sparse terminal network and many large cells, which might represent motoneurons. A sparse network of varicose cholecystokinin-immunoreactive fibers was found evenly distributed in the grafts. In normal spinal cord a dense cholecystokinin-positive network of primary sensory afferent origin was found in the dorsal horn. In the grafts cholecystokinin cell bodies were seen after colchicine treatment.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗