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The nervus terminalis of the guinea pig: a new luteinizing hormone-releasing hormone (LHRH) neuronal system.

Immunoreactive LHRH-like material has been found in the cells and fibers of the nervus terminalis in fetal and adult guinea pig brains. LHRH-containing neurons and axons are seen in the nasal mucosa intermingled with fibers of the olfactory nerves, in ganglia along the ventromedial surfaces of the olfactory bulbs and forebrain, and in clusters surrounding perforating branches of the anterior cerebral artery in the regions of the septal nuclei and olfactory tubercle. Nonreactive neurons are found adjacent to the LHRH-positive cells in all of the ganglia. LHRH-immunoreactive cells and axons of the nervus terminalis are in intimate contact with cerebral blood vessels and the cerebrospinal fluid along the intracranial course of this nerve, deep to the meninges. The possible involvement of these structures in the neural mechanisms of sexual behavior and the neurohormonal regulation of reproductive function are discussed.

Age Factors↗

Ontogenic origin of salmon GnRH neurons in the ventral telencephalon and the preoptic area in masu salmon.

During the ontogeny of masu salmon Oncorhynchus masou, neurons producing the salmon type of gonadotropin-releasing hormone (sGnRH) were first detected in the olfactory epithelium of the eyed egg and, subsequently, in the brain, suggesting a migration of these cells. Among sGnRH neurons distributed from the olfactory nerve (ON) through the preoptic area (POA), those in the ventral telencephalon (VT) and the POA are indicated to regulate gonadotropin secretion. Thus, it is of interest to know whether all the sGnRH neurons originate from the olfactory epithelium. In the present study, we examined by in situ hybridization whether sGnRH neurons are present in the VT-POA of fish, whose olfactory epithelia including sGnRH clusters were cauterized just after hatching (44 days after fertilization). Fish were sampled in June (212 days after the operation). Neurons expressing sGnRH mRNA were detected in the VT-POA as well as in the ON, ventral olfactory bulb, and transitional area between the olfactory bulb and telencephalon (which is considered to correspond to the terminal nerve ganglion) in the control group. In contrast, neurons expressing sGnRH mRNA were not detected in the VT-POA in the olfactory epithelium lesioned (OEL) group. Furthermore, pituitary sGnRH content in the OEL group was just above the detectable limit and was significantly lower than that in the corresponding control group in both sexes. These results indicate that sGnRH neurons in the VT-POA are derived from the olfactory epithelium in masu salmon, although the possibility cannot be ruled out that sGnRH neurons in the VT-POA arise from the VT-POA, but were delayed in expressing sGnRH because of the trauma of cauterization.

Animals↗

Ontogenetic organization of the FMRFamide immunoreactivity in the nervus terminalis of the lungfish, Neoceratodus forsteri.

The development of the nervus terminalis system in the lungfish, Neoceratodus forsteri, was investigated by using FMRFamide as a marker. FMRFamide immunoreactivity appears first within the brain, in the dorsal hypothalamus at a stage around hatching. At a slightly later stage, immunoreactivity appears in the olfactory mucosa. These immunoreactive cells move outside the olfactory organ to form the ganglion of the nervus terminalis. Immunoreactive processes emerge from the ganglion of the nervus terminalis in two directions, one which joins the olfactory nerve to travel to the brain and the other which courses below the brain to enter at the level of the preoptic nucleus. Neither the ganglion of the nervus terminalis nor the two branches of the nervus terminalis form after surgical removal of the olfactory placode at a stage before the development of FMRFamide immunoreactivity external to the brain. Because this study has confirmed that the nervus terminalis in lungfish comprises both an anterior and a posterior branch, it forms the basis for discussion of homology between these branches and the nervus terminalis of other anamniote vertebrates.

Animals↗

Expression of intermediate filaments and desmoplakin in vertebrate olfactory mucosa.

The expression of intermediate filaments (IF) and desmoplakin was investigated in frog, bovine, and human (fetal) olfactory mucosa. IF are tissue-specific molecular cytoskeletal markers; desmoplakin is the major desmosomal protein. Positive immunoreactivity was observed in the epithelium and in the subepithelial Bowman's glands to keratin and to desmoplakin, indicating the epithelial nature of this tissue. Desmin, neurofilaments, and glial fibrillary acidic protein (GFAP) were not detected in the mucosa. The absence of neurofilaments and GFAP in the tissue containing sensory neurons and glia-like supporting cells is a unique feature and may be related to the fact that the chemosensory neurons are situated in a bonafide epithelium and are known to undergo continuous turnover. In view of the controversy regarding the expression of vimentin in the olfactory neurons, three independently derived antibodies to vimentin were used; weak or no labeling was found in the epithelium, whereas mesenchymal cells in the lamina propia were labeled with all three antibodies. Olfactory nerve fascicles in the lamina propia were heterogenously labeled: VIM 13.2 gave very weak labeling; aVimAS showed mild labeling and SBV-21 showed intensive labeling in the nerve fascicle. This heterogenous labeling pattern may suggest that olfactory vimentin is distinct in reacting only with some of the antivimentin antibodies.

Animals↗

Pharmacological analysis of slow potentials recorded in frog olfactory bulb during natural stimulation.

Pharmacological agents (strychnine, picrotoxin, pentobarbital, chloralose, GABA, penicillin, morphine) were used to investigate the nature of the slow potential recorded in the frog olfactory bulb in response to natural stimulation. Three possible hypotheses were tested: 1) The slow potential is neuroglial in nature; 2) it is the analog of the dorsal-root potential of the spinal cord and reflects depolarization of primary afferents arising in the terminals of the olfactory nerve and responsible for presynaptic inhibition in the frog olfactory bulb; 3) the slow potential reflects postsynaptic processes. The results showed great similarity between changes in the slow and dorsal-root potentials of the spinal cord in response to the action of pharmacological agents. However, the slow potential is evidently a complex response and incorporates at least one other component - depolarization of the dendrites of unknown nature.

Action Potentials↗

Scanning electron microscopic study of degeneration and regeneration in the olfactory epithelium after axotomy.

The olfactory epithelium of the adult hamster (Mesocricetus auratus) was examined with the scanning electron microscope following olfactory nerve axotomy. Axotomy results in retrograde degeneration of mature olfactory neurons. Maximum degeneration was observed around day 4. During the degeneration period the epithelium consists primarily of supporting and basal cells. Microvillar columnar supporting cells were observed to have fine cellular processes extending from their lateral border to neighbouring cells. Supporting cells extended to the basal lamina where they terminated in foot-like processes of variable shapes (club, splay and hook). Basal cells which gave rise to new replacement olfactory neurons were observed near the basal lamina. They had a rough cellular surface covered with small granules and fine cellular extensions. Bowman's gland duct cells extended unbranched through the epithelium where they formed funnel duct openings covered with microvilli. During early recovery periods (5-30 days) the number of olfactory neurons in the lower epithelium region increased. We observed olfactory neurons with developing axon and dendritic processes. Specialized growth cone structures were seen at the tips. Olfactory neuron growth cones were elongated or club-shaped and had a ruffled membrane surface. Several thin filopodia extended from the growth cone and made contact with adjacent cells. At late recovery periods (35-120 days) there was a marked increase in the number of olfactory neurons within the middle and lower epithelium regions. Numerous dendritic processes extended to the epithelial surface and terminated in knob-like ciliated structures. Olfactory axons passed basally, forming small intra-epithelial bundles that penetrated the basal lamina then fasciculated into larger bundles within the lamina propria. This study provides detailed three-dimensional observations of the olfactory epithelium following neuron injury, and describes neural degenerative changes, replacement of olfactory neurons, development and maturation. In addition, we describe the structure and basal attachment of supporting cells and their glial-like relation with olfactory neurons.

Animals↗

Developmental brain abnormalities accompanied with the retarded production of S-100 beta protein in genetic polydactyly mice.

The homozygotes of a mouse strain with genetic polydactyly (Polydactyly Nagoya, Pdn) exhibit various brain malformations including exencephaly in about 20%. In the present report, the brains of homozygotes (Pdn/Pdn) which were not exencephalic were examined morphologically and biochemically. Homozygous newborn brains showed hydrocephaly, some gyri on the cerebral hemisphere, absence of the corpus callosum, absence of the commissura anterior, absence of the fornix and commissura fornicis, protuberance of the cortical tissue from the brain surface, and abnormal architecture of the hippocampus. An irregular mass of olfactory nerve was observed on the cribriform plate, and the olfactory bulb was deficient. From these findings, we considered Pdn/Pdn as a kind of arhinencephalic mouse. Nervous tissue-related proteins, S-100 alpha, S-100 beta, creatine kinase B (CK-B), neuron-specific gamma-enolase, guanosine triphosphate binding proteins (Go alpha, Gi2 alpha and G beta) were immunoassayed in the cerebrum of Pdn/Pdn embryos and newborns. Among the protein analysed, only S-100 beta of Pdn/Pdn showed a significantly lower level than that of +/+ cerebrum during the observation period. The newborn brains were examined immunohistochemically using S-100 alpha, S-100 beta, CK-B, Go alpha and NSE antibodies. We could find no differences in the staining patterns among the Pdn/Pdn, Pdn/+ and +/+ brains.

Animals↗

Developmental expression of three different prepro-GnRH (gonadotrophin-releasing hormone) messengers in the brain of the European sea bass (Dicentrarchus labrax).

In this study, we have analyzed the ontogenic expression of three gonadotrophin-releasing hormones (GnRH) systems expressed in the brain of a perciform fish, the European sea bass, using in situ hybridization. The riboprobes used correspond to the GnRH-associated peptide (GAP) coding regions of the three prepro-GnRH cDNAs cloned from the same species: prepro-salmon GnRH, prepro-seabream GnRH and prepro-chicken GnRH II. On day 4 after hatching, the first prepro-chicken GnRH-II mRNA-expressing cells appeared in the germinal zone of the third ventricle. They increased in number and size from 10 to 21 days, reaching at day 30 their adult final position, within the synencephalic area, at the transitional zone between the diencephalon and the mesencephalon. First prepro-salmon GnRH mRNA-expressing cells became evident on day 7 arising from the olfactory placode and migrating towards the olfactory nerve. On day 10, this cell group reached the olfactory bulb, being evident in the ventral telencephalon and preoptic area from days 15 and 45, respectively. Weakly labeled prepro-seabream GnRH mRNA-expressing cells were first detected at 30 days in the olfactory area and ventral telencephalon. On day 45, prepro-seabream GnRH mRNA-expressing cells were also present in the preoptic region reaching the ventrolateral hypothalamus on day 60. The results obtained in sea bass indicate that sGnRH and sbGnRH cells have a common origin in an olfactory primordium suggesting that both forms might arise from a duplication of a single ancestral gene, while cGnRH-II cells develop from a synencephalic primordium.

Animals↗

Presynaptic afferent inhibition of lobster olfactory receptor cells: reduced action-potential propagation into axon terminals.

Action-potential propagation into the axon terminals of olfactory receptor cells was measured with the use of voltage-sensitive dye imaging in the isolated spiny lobster brain. Conditioning shocks to the olfactory nerve, known to cause long-lasting suppression of olfactory lobe neurons, allowed the selective imaging of activity in receptor cell axon terminals. In normal saline the optical signal from axon terminals evoked by a test stimulus was brief (40 ms) and small in amplitude. In the presence of low-Ca2+/high-Mg2+ saline designed to reduce synaptic transmission, the test response was unchanged in time course but increased significantly in amplitude (57 +/- 16%, means +/- SE). This increase suggests that propagation into receptor cell axon terminals is normally suppressed after a conditioning shock; this suppression is presumably synaptically mediated. Thus our results show that presynaptic inhibition occurs at the first synapse in the olfactory pathway and that the inhibition is mediated, at least in part, via suppression of action-potential propagation into the presynaptic terminal.

Action Potentials↗

[The effect of temperature changes on evoked potentials of the fish olfactory bulb].

Perches were kept at 10 degrees C and fixed in a chamber through which water was perfused. During the experiment the water temperature was changed. The potentials induced by electric stimulation of the olfactory nerve were recorded from the dorsal surface of the olfactory bulb. The oscillations of the evoked potentials separated into several components when the temperature was lowered. Six components were found in the evoked response of the olfactory bulb at 8 degrees C.

Animals↗

[Spatio-temporal study of the electrical activities of the olfactory bulb of the trout Salmo gairdneri].

The electrical stimulation of the olfactory nerve allows to record steady field potentials in the olfactory bulb of the trout Salmo gairdneri. 200 microns separate recording points which are localised on a tridimensional network. At 18, 24, 30 and 38 ms the amplitudes are measured and plotted in a calculator which determines the isopotentials in any vertical, horizontal and sagittal planes. With this method, we have an instantaneous and global vision of bulbar activity and we observe striking anatomo-functional correlations in the olfactory bulb of Salmo gairdneri.

Animals↗

Expression of hepatocyte growth factor/scatter factor, its receptor, c-met, and tissue-type plasminogen activator during development of the murine olfactory system.

The expression of hepatocyte growth factor/scatter factor (HGF/SF) and its receptor, the c-met proto-oncogene product, was examined by in situ hybridization in the developing and adult murine olfactory system and compared with the expression of a known activator of HGF/SF, tissue-type plasminogen activator (tPA). In the developing olfactory canal, expression of both c-met and tPA was observed in the olfactory neuroepithelium, whereas HGF/SF expression appeared to be confined to the mucosa adjacent to the neuroepithelium. During development of the olfactory bulb, HGF/SF and tPA were expressed within the rostral migratory pathway leading to the olfactory bulb, whereas c-met expression was observed in the mitral cell layer (MCL) of the olfactory bulb and in the anterior olfactory nucleus. In the adult olfactory bulb, expression of HGF/SF was restricted to the periglomerular region of the glomerular layer, whereas c-met was expressed in the MCL and olfactory nerve fiber layers (ONL). tPA expression in the adult olfactory bulb was observed in the ONL, MCL, and granule cell layers. Therefore, tPA expression was relatively coincident with the expression of HGF/SF and/or c-met in the appropriate projection patterns of the developing and adult olfactory system. In addition, antibodies against tPA inhibited the olfactory bulb extract-mediated cleavage of single-chain HGF/SF. These results suggest that tPA may play a regulatory role in the development and maintenance of the olfactory system by activating HGF/SF in the immediate vicinity of its receptor.

Age Factors↗

Role of neuropeptide Y in the regulation of gonadotropin releasing hormone system in the forebrain of Clarias batrachus (Linn.): immunocytochemistry and high performance liquid chromatography-electrospray ionization-mass spectrometric analysis.

Although the importance of neuropeptide Y (NPY) in the regulation of gonadotropin releasing hormone (GnRH) and reproduction has been highlighted in recent years, the neuroanatomical substrate within which these substances might interact has not been fully elucidated. Present work was undertaken with a view to define the anatomical-physiological correlates underlying the role exercised by NPY in the regulation of GnRH in the forebrain of the teleost Clarias batrachus. Application of double immunocytochemistry revealed close associations as well as colocalizations of the two peptides in the olfactory receptor neurons (ORNs), olfactory nerve fibers and their terminals in the glomeruli, ganglion cells of nervus terminalis, medial olfactory tract, fibers in the area ventralis telencephali/pars supracommissuralis and cells as well as fibers in the pituitary. NPY containing axons were found to terminate in the vicinity of GnRH cells in the pituitary with light as well as electron microscopy. Double immunoelectron microscopy demonstrated gold particles for NPY and GnRH colocalized on the membrane and in dense core of the secretory granules in the cells distributed in all components of the pituitary gland. To assess the physiological implication of these observations, NPY was injected via the intracranial route and the response of GnRH immunoreactive system was evaluated by relative quantitative morphometry as well as high performance liquid chromatography (HPLC) analysis. Two hours following NPY (20 ng/g body weight) administration, a dramatic increase was observed in the GnRH immunoreactivity in the ORNs, in the fibers of the olfactory bulb (163%) and medial olfactory tract (351%). High performance liquid chromatography-electrospray ionization-mass spectrometric analysis confirmed the immunocytochemical data. Significant rise in the salmon GnRH (sGnRH)-like peptide content was observed in the olfactory organ (194.23%), olfactory bulb (146.64%), telencephalon+preoptic area (214.10%) and the pituitary (136.72%) of the NPY-treated fish. However, GnRH in the hypothalamus was below detection limit in the control as well as NPY-treated fish. Present results suggest the involvement of NPY in the up-regulation of sGnRH containing system at different level of neuraxis extending from the olfactory epithelium to the pituitary in the forebrain of C. batrachus.

Animals↗

A disintegrin and metalloprotease 21 (ADAM21) is associated with neurogenesis and axonal growth in developing and adult rodent CNS.

We have reported that alpha6beta1 integrin regulates the directed migration of neuroblasts from the adult rodent subventricular zone (SVZ) through the rostral migratory stream (RMS). ADAM (a disintegrin and metalloprotease) proteins bind integrins. Here, we show that ADAM21, but not ADAM2, -3, -9, -10, -12, -15, or -17, is expressed in adult rats and mice by ependyma and SVZ cells with long basal processes, and in radial glia at early postnatal times. ADAM21-positive processes projected into the RMS, contacted blood vessels, and were present within the RMS intermingled with neuroblasts up to where neuroblasts start their radial migration and differentiation in the olfactory bulb. Tissue inhibitors of metalloproteases (TIMPs) 1, 2, and 3 are present in the ependymal layer but not in the SVZ and RMS. Thus, ADAM21 could regulate neurogenesis and guide neuroblast migration through cleavage-dependent activation of proteins and integrin binding. ADAM21 is also present in growing axonal tracts during postnatal development and in growing primary olfactory axons in adults. In the olfactory nerve layer, ADAM21 often, but not always, colocalizes with OMP, a marker of mature olfactory neurons, but is not colocalized with the immature marker betaIII-tubulin. This suggests that ADAM21 is involved in the final axonal outgrowth phase and/or synapse formation. TIMP3 is present in periglomerular neurons, where it could restrict ADAM21-mediated axonal growth to the glomeruli. ADAM21's unique disintegrin and metalloprotease sequences and its restricted expression suggest that it might be a good target for influencing neurogenesis and neuronal plasticity.

ADAM Proteins↗

Responsiveness of the olfactory receptor cells in dog to some odors.

A preparation has been developed in the dog which allows recording the electrical activity from an olfactory nerve twig containing the axons of a small group of olfactory receptor cells. The dog's response to n-pentyl acetate is vigorous and stable, like that of other air-breathing animals. The dog's response magnitude dependence on the nasal flow rate was noticeable for n-pentyl acetates, but not so great as for n-butyric acid. The response to n-butyric acid strongly depends on the nasal flow. The start of the nasal air flow caused an increase of neural activity, which is called flow response. The results show that the nasal flow rate is a very important factor which determines the response to odors. Methyl p-hydroxybenzoate is known as a dog's pheromone, however, this odor caused the feeble response in the electrical activity of the dog's olfactory receptor cells. The differences may be dependent on several factors.

Animals↗

Expression of non-N-methyl-D-aspartate glutamate receptor subunits in the olfactory epithelium.

The channel properties of the multimeric ionotropic glutamate receptors can be regulated by their subunit composition. The relationship between the structure and physiological functions of glutamate receptors, however, is difficult to study in the CNS because of the large number of these subunits, their widespread distribution, and neuronal heterogeneity. To avoid these difficulties, and to uncover possible novel functions of ionotropic glutamate receptors in sensory neurons, we examined the expression of non-N-methyl-D-aspartate glutamate receptor subunits in a simple neuronal system: the olfactory epithelium. It contains only one neuronal type, the olfactory receptor neuron, that receives no synaptic innervation within the epithelium and therefore should not require conventional postsynaptic glutamate receptors. The axons of these neurons, however, terminate and release glutamate in the glomerular region of the olfactory bulb, and may contain presynaptic glutamate receptors. By reverse transcriptase-polymerase chain reaction amplification and RNase protection assays, we showed that a subset of non-N-methyl-D-aspartate receptor subunits is expressed in the olfactory epithelium. The most abundant is KA2, which can form kainate-selective ion channels with GluR5 or GluR6. Messenger RNAs for GluR6, and for the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate/kainate-type (AMPA/KA) GluR2 and GluR3 subunits, are also present, but at levels lower than that of KA2 by an order of magnitude. In situ hybridization and immunocytochemistry localized KA2 to only the olfactory receptor neurons, and not to any other cell type in the olfactory epithelium. Surprisingly, antibodies against KA2 or GluR5/6/7 primarily stained the olfactory neuron dendritic knobs that are specialized for odorant signalling at the sensory epithelial lumenal surface, and the olfactory neuron axon bundles that project to the olfactory bulb. The presence of a limited subset of non-N-methyl-D-aspartate receptor subunits in the olfactory epithelium, and the localization of a kainate-selective receptor to both the axons and specialized dendritic knobs of olfactory receptor neurons, which receive no known synaptic input, suggest that these non-N-methyl-D-aspartate receptor subtypes may mediate either novel non-synaptic functions in the olfactory neuron dendrites or presynaptic functions in the olfactory nerve terminals or axons. These data also suggest that the olfactory sensory system, possessing a relatively simple anatomical organization and a limited number of glutamate receptor subunits, may be useful for elucidating facets of the complex relationships between subunit composition and physiological function of ionotropic glutamate receptors.

Animals↗

Olfactory marker protein (OMP) gene deletion causes altered physiological activity of olfactory sensory neurons.

Olfactory marker protein (OMP) is an abundant, phylogentically conserved, cytoplasmic protein of unknown function expressed almost exclusively in mature olfactory sensory neurons. To address its function, we generated OMP-deficient mice by gene targeting in embryonic stem cells. We report that these OMP-null mice are compromised in their ability to respond to odor stimull, providing insight to OMP function. The maximal electroolfactogram response of the olfactory neuroepithelium to several odorants was 20-40% smaller in the mutants compared with controls. In addition, the onset and recovery kinetics following isoamyl acetate stimulation are prolonged in the null mice. Furthermore, the ability of the mutants to respond to the second odor pulse of a pair is impaired, over a range of concentrations, compared with controls. These results imply that neural activity directed toward the olfactory bulb is also reduced. The bulbar phenotype observed in the OMP-null mouse is consistent with this hypothesis. Bulbar activity of tyrosine hydroxylase, the rate limiting enzyme of catecholamine biosynthesis, and content of the neuropeptide cholecystokinin are reduced by 65% and 50%, respectively. This similarity to postsynaptic changes in gene expression induced by peripheral olfactory deafferentation or naris blockade confirms that functional neural activity is reduced in both the olfactory neuroepithelium and the olfactory nerve projection to the bulb in the OMP-null mouse. These observations provide strong support for the conclusion that OMP is a novel modulatory component of the odor detection/signal transduction cascade.

Animals↗

Olfactory ensheathing cells: bridging the gap in spinal cord injury.

Spinal cord injury (SCI) continues to be an insidious and challenging problem for scientists and clinicians. Recent neuroscientific advances have changed the pessimistic notion that axons are not capable of significant extension after transection. The challenges of recovering from SCI have been broadly divided into four areas: 1) cell survival; 2) axon regeneration (growth); 3) correct targeting by growing axons; and 4) establishment of correct and functional synaptic appositions. After acute SCI, there seems to be a therapeutic window of opportunity within which the devastating consequences of the secondary injury can be ameliorated. This is supported by several observations in which apoptotic glial cells have been identified up to 1 week after acute SCI. Moreover, autopsy studies have identified anatomically preserved but unmyelinated axons that could potentially subserve normal physiological properties. These observations suggest that therapeutic strategies after SCI can be directed into two broad modalities: 1) prevention or amelioration of the secondary injury, and 2) restorative or regenerative interventions. Intraspinal transplants have been used after SCI as a means for restoring the severed neuraxis. Fetal cell transplants and, more recently, progenitor cells have been used to restore intraspinal circuitry or to serve as relay for damaged axons. In an attempt to remyelinate anatomically preserved but physiologically disrupted axons, newer therapeutic interventions have incorporated the transplantation of myelinating cells, such as Schwann cells, oligodendrocytes, and olfactory ensheathing cells. Of these cells, the olfactory ensheathing cells have become a more favorable candidate for extensive remyelination and axonal regeneration. Olfactory ensheathing cells are found along the full length of the olfactory nerve, from the basal lamina of the epithelium to the olfactory bulb, crossing the peripheral nervous system-central nervous system junction. In vitro, these cells promote robust axonal growth, in part through cell adhesion molecules and possibly by secretion of neurotrophic growth factors that support axonal elongation and extension. In animal models of SCI, transplantation of ensheathing cells supports axonal remyelination and extensive migration throughout the length of the spinal cord. Although the specific properties of these cells that govern enhanced axon regeneration remain to be elucidated, it seems certain that they will contribute to the establishment of new horizons in SCI research.

Acute Disease↗