Distribution of NT-IR perikarya in the brain of the guinea pig with special reference to cardiovascular centers in the medulla oblongata

The occurrence and distribution of neurotensin-immunoreactive (NT-IR) perikarya was studied in the central nervous system of the guinea pig using a newly raised antibody (KN 1). Numerous NT-IR perikarya were found in the nuclei amygdaloidei, nuclei septi interventriculare, hypothalamus, nucleus parafascicularis thalami, substantia grisea centralis mesencephali, ventral medulla oblongata, nucleus solitarius and spinal cord. The distribution of NT-IR perikarya was similar to that previously described in the rat and monkey. In the gyrus cinguli, hippocampus and nucleus olfactorius, though, no NT-IR neurons were detected in this investigation. Additional immunoreactive perikarya, however, were observed in areas of the ventral medulla oblongata, namely in the nucleus paragigantocellularis, nucleus retrofacialis and nucleus raphe obscurus. The relevance of the NT-IR perikarya within the ventral medulla oblongata is discussed with respect to other neuropeptides, which are found in this area, and to cardiovascular regulation.     

Cytochemical localization of alkaline phosphatase in the ependyma of the rat medulla oblongata

Summary The ependyma of the IVth ventricle and the central canal of the rat medulla oblongata was investigated using the cytochemical technique for alkaline phosphatase (AlPase) which revealed two types of ependymal cells in the medulla. The central canal type of the ependymal cell occupying the dorsal part of the central canal in the lower medulla exhibited intense AlPase activity with light microscopy. These cells had reaction products in all plasma membranes, including the microvilli and the cilia at the luminal cell surface. Some cells appeared to be tanycytes, since the process reached the basement membrane of the parenchymal blood vessel. The ventricular type of ependymal cells, which form the floor of the IVth ventricle and the central canal, contained no reaction products in any structure of the luminal cell surface.The possible relationship between the cerebrospinal fluid and the nervous tissues through the ependymal linings is discussed.     

Respiratory effects of H+ and dinitrophenol injections into the brain stem subarachnoid space of fetal lambs.

Mock cerebrospinal fluid (pH 5.37-8.38) or 2,4-dinitrophenol (DNP) (0.15-1.5 mg) was injected into the subarachnoid space of the ventral brain stem of exteriorized fetal sheep. Changes in pH on the ventral surface of the medulla did not stimulate respiratory efforts or induce significant cardiovascular changes. The respiratory response to DNP injections ranged from no response to prolonged rhythmic ventilation that was independent of the peripheral chemoreceptors or the control arterial pH and blood gas tensions. This inconsistency suggests an effector site somewhat removed from the immediate surface of the medulla. The heart rate and blood pressure were not affected. It is concluded that increased H+ concentration in the extracellular fluid of the fetal ventral medulla does not initiate respiration, and any respiratory response to metabolic inhibitors applied to this area therefore is not attributable to a secondary change in surface pH.     

Identification of a subsurface area in the ventral medulla sensitive to local changes in PCO2.

The exact location of the central respiratory chemoreceptors sensitive to changes in PCO2 has not yet been determined. To avoid the confounding effects of the cerebral circulation, we used the in vitro brain stem-spinal cord of neonatal rats (1-5 days old) to identify areas within 500 microns of the ventral surface of the medulla where changes in PCO2 evoked a sudden increase in the rate of respiratory neural activity. The preparation was superfused with mock cerebrospinal fluid (CSF) while maintained at constant temperature (26 +/- 1 degrees C) and pH (7.34). Respiratory frequency increased linearly with decreases in superfusate pH (r2 = 0.92, P less than 0.001), indicating that the respiratory circuitry for the detection of CO2 and stimulation of breathing was intact in this preparation. The search for central chemoreceptors was performed with a specially designed micropipette that allowed microejection of 2-10 nl of mock CSF equilibrated with different CO2-O2 gas mixtures. The pipette was advanced in 50- to 100-microns steps by use of a microdrive to a maximum depth of 500 microns from the surface of the ventral medulla. Depending on the location of the micropipette, ejection of CO2-acidified mock CSF at depths of 100-350 microns below the ventral surface of the medulla stimulated neural respiratory output. Using this response as an indication of the location of central respiratory chemoreceptors, we found that chemoreceptive elements were located in a column in the ventromedial medulla extending from the hypoglossal rootlets caudally to an area 0.75 mm caudal to VI nerve in the rostral medulla.(ABSTRACT TRUNCATED AT 250 WORDS)     

Substance P in the ventrolateral medulla oblongata regulates ventilatory responses.

Local injection of substance P (SP) into the ventral portion of the nucleus gigantocellularis, nucleus reticularis lateralis, and nucleus retrofacialis of the ventrolateral medulla oblongata (VLM) or direct application on the ventral surface of the medulla oblongata caused marked stimulation of tidal volume (VT) and/or minute ventilation (VE). The ventilatory response to hypoxia was significantly blunted after SP in the VLM but not in the dorsal medulla oblongata (DM) (nucleus tractus solitarius). The SP antagonist [D-Pro2,D-Trp7,9]SP almost completely inhibited this response when applied locally to a wide area of the superficial layer of the VLM but not of the DM. Unilateral or bilateral application of 0.3-1.5 nmol of the SP antagonist in the VLM (corpus trapezoideum and the caudal region extending from the rootlets of the nucleus hypoglossus to the first cervical segment) markedly attenuated the response to a 5% CO2 inhalation. The inhibition of the CO2 response was seen after [D-Pro2,D-Trp7,9]SP in the rostral areas of the medulla oblongata corresponding to the corpus trapezoideum and the caudal region extending from the rootlets of the nucleus hypoglossus to the first cervical segment of the cervical cord. Electric somatosensory-induced ventilatory stimulation could be depressed by approximately 70% by [D-Pro2,D-Trp7,9]SP locally applied on the surface of the VLM. We conclude that SP is involved in the hypoxic, hypercapnic, and somatosensory ventilatory responses in the rat. However, these respiratory reflexes are mediated via different neuronal pools in the medulla oblongata, mainly the VLM.     

Peptide YY-like immunoreactivity in the central nervous system of the rat

The concentration of peptide YY (PYY)-like immunoreactivity in rat brain and spinal cord was determined by radioimmunoassay. The highest concentrations were found in the cervical spinal cord (18.1 +/- 1.3 ng/g, mean +/- S.E.M.) and in the medulla oblongata (16.3 +/- 1.5 ng/g). Lower amounts were found in the pons and in the hypothalamus. Chromatographic analysis of the PYY-like immunoreactivity from various regions of the brain revealed 95% of the immunoreactive material to be indistinguishable from synthetic porcine PYY. PYY-immunoreactive nerve cell bodies could be demonstrated by immunocytochemistry in the medulla oblongata of colchicine-treated rats, the largest group of cells being found in the midline area between and partly in the raphe pontis and obscurus nuclei. Another large group of immunoreactive cells was detected more laterally in the medial parts of the gigantocellular reticular nucleus. A few cells, finally, were seen in the dorsal parts of the medulla, including the nucleus of the solitary tract. Varicose nerve fibers displaying PYY immunoreactivity were observed in many parts of the hypothalamus, pons, medulla and spinal cord.     

Localization of three types of arginine vasotocin receptors in the brain and pituitary of the newt Cynops pyrrhogaster

The distribution of three types of arginine vasotocin (AVT) receptors in the brain and pituitary of the newt Cynops pyrrhogaster, namely, the V1a-, V2-, and V3/V1b-type receptors, was studied by means of in situ hybridization and immunohistochemistry. mRNA signals and immunoreactive cells for the V1a-type receptor were observed in the telencephalon (mitral layer of the olfactory bulb, dorsal and medial pallium, lateral and medial amygdala, bed nucleus of the decussation of the fasciculus telencephali, bed nucleus of the stria terminalis), diencephalon (anterior preoptic area, magnocellular preoptic nucleus, suprachiasmatic nucleus, ventral thalamus, dorsal and ventral hypothalamic nucleus), mesencephalon (tegmentum, interpeduncular nucleus), and medulla oblongata (median reticular formation, nucleus motorius tegmenti). Cells expressing the V2-type receptor were found in the telencephalon (medial pallium, lateral and medial amygdala, bed nucleus of the decussation of the fasciculus telencephali), and mesencephalon (tegmentum trigemini and facialis). In the paraphysis (possibly the main site of cerebrospinal fluid production), only V2-type receptor mRNA signal and immunoreactivity were detected. V3/V1b-type receptor mRNA was expressed in the diencephalon (dorsal hypothalamic nucleus, nucleus tuberculi posterioris), mesencephalon (tegmentum, interpeduncular nucleus), and medulla oblongata (raphe nucleus), whereas V3/V1b-type-receptor-like immunoreactivity was scarcely detectable in the entire brain. The V3/V1b-type receptor was predominantly expressed in the anterior pituitary. V3/V1b-type receptor and proopiomelanocortin mRNAs were co-localized in the distal lobe of the pituitary. This is the first report of the distribution of three types of AVT receptor in the brain and pituitary of non-mammalian vertebrates.     

Cell junctions in the subcommissural organ of the rabbit as revealed by use of ruthenium red

Summary Tight junctions were found in the apical junctional complex of the adult rabbit subcommissural organ (SCO) in addition to zonulae adhaerentes and gap junctions of typical ependymal cells. Ventricular perfusion of ruthenium red before fixation was found to give excellent results for distinguishing between gap and tight junctions at the ependymal surface. The implication of tight junctions as a mechanical means of sealing off the SCO area from the cerebrospinal fluid and the use of ruthenium red as a tracer substance are discussed.This work was supported by grants from Statens almindelige Videnskabsfond, Copenhagen.     

History and status of the area postrema

The history of morphological and functional studies on the area postrema (AP) is traced for significant landmarks from 1896, when its name was conferred by Retzius, to 1960 when the foundation of inquiry had become firmly set. A comparative anatomical survey of the medulla oblongata identifies the AP in mammals and birds but not in amphibians and lower phyla even though other members of the so-called circumventricular organ system are represented in the more ancient creatures. Existence of the AP in reptiles is insufficiently documented. The transition from water- to land-dwelling animal life affords propitious neural remodeling for the emergence of the AP in evolving species. No vital role is known to be served by the AP. Nonetheless, its physical interposition between the blood and cerebrospinal fluid and its shared functions with the nucleus tractus solitarii indicate a capability for widespread somatovisceral influence in response to particular perturbations. It is suggested that the diverse systemic expressions of postremal activation are encompassed in the general syndrome of nausea and vomiting.     

大鼠延髓尾端加压区对外周血管紧张的影响

本文以血管的灌流压变化反映血管紧张性,观察Ｌ－谷氨酸及肾上腺素能受体阻断剂微量注射于大鼠延髓腹面心血管活动调控区对骨骼肌血管和肾血管紧张的影响。实验观察到,Ｌ－－ｇｌｕ兴奋延髓尾端加压区（ＣＰＡ）在动脉直升的同时,双后肢骨骼肌灌流压上升３７．５９％,左肾灌流压上升１７．０４％,如预先在延髓腹面加我注入普萘少尔或酚妥拉明均能明显减小Ｌ－ｇｌｕ兴奋ＣＰＡ时的灌流压和肾灌流压升高效应。提示ＣＰＡ对外周血     

Chemosensitive structures of the medulla oblongata participate in blood pressure regulation and belong to APUD-system

It was shown in 25 experiments on anesthetized spontaneously breathing and immobilized artificially ventilated cats that the cooling of the intermediate chemosensitive area of the ventral surface of medulla oblongata to 20-22 degrees C is followed by depressor reaction that is more pronounced after preliminary occlusion of the sinocarotid and vagus nerves. Two types of apudocytes were revealed in this brain area with the use of the histochemical and immunohistochemical techniques. The first type cells are typically nervous and contain noradrenaline. The second type apudocytes adjacent to the microvascular wall showed melatonin and its precursors. It is assumed that the cells detected participate in the arterial pressure control.     

Developmental study of the shortened spinal cord in the adult tiger puffer fish, Takifugu rubripes (Teleostei)

ABSTRACT The spinal cords of vertebrates are generally divided into the cord proper and the minute filum terminale. While the spinal cord extends the entire length of the vertebral canal in the adult tiger puffer, Takifugu rubripes, the cord proper is greatly reduced in length and almost all of the canal is occupied by the filum terminale, which is tape-like rather than thread-like. The dorsal and ventral roots of the spinal nerves extend, respectively, above and below the filum terminale; as a whole, these form a massive cauda equina. Supramedullary cells are found in the rostral half of the medulla oblongata caudal to the cerebellum. In 4-mm long tiger puffers, the spinal cord is cylindrical and supramedullary cells are found in the rostral half of the cord. In 7-mm puffers, the longitudinally arranged ventral roots appear ventrally in the middle portion of the spinal cord. In 15-mm puffers, the dorsal and ventral roots run longitudinally along the spinal cord and have noticeably increased in number. Supramedullary cells are located in the rostral 15% of the cord. In 21-mm puffers, the spinal cord in large part becomes dorsoventrally flattened. In 30-mm puffers, the spinal cord becomes much flatter, and supramedullary cells now are located mainly in the medulla oblongata. These observations indicate that formation of the shortened spinal cord proper is due to at least two developmental processes. First, the elongation of the spinal cord proper is remarkably less than that of the vertebral canal. Second, the bulk of the spinal cord proper is translocated to the cranial cavity, where it is transformed into part of the medulla oblongata.     

Immunohistochemical localization of neuropeptide Y-like substance in the brain and hypophysis of the brown hagfish,Paramyxine atami

The distribution of neuropeptide Y-like immunoreactivity in the brain and hypophysis of the brown hagfish, Paramyxine atami, was examined by use of the peroxidase-antiperoxidase method. Immunoreactive cells were found in two areas of the brain, the nucleus hypothalamicus of the diencephalon and the ventrolateral area of the caudal tegmentum, at the level of the nucleus motorius V–VII. The labeled cells of the nucleus hypothalamicus were loosely grouped and recognized as bipolar neurons. Immunolabeled fibers were widely distributed in the brain, showing the highest density in the diencephalon. They were sparse, or absent, in the olfactory bulb, habenula, primordium hippocampi, neurohypophysis, corpus interpedunculare, and dorsolateral area of the medulla oblongata. The fibers appeared to project exclusively from the ventral hypothalamus to various other portions of the brain: the anterolateral areas of the telencephalon via the basal hypothalamus, the pars dorsalis thalami, the dorsocaudal region of the mesencephalon, and the ventromedial portions of the tegmentum and anterior medulla oblongata. These findings suggest that, in the brown hagfish, NPY-like substance is involved in neuroregulation of various cerebral areas, but it may be of little significance in the control of pituitary function.     

Distribution of choline acetyltransferase and acetylcholinesterase in the nervous system of the dog

Choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) activity were determined in 23 selected parts of the dog CNS and 4 parts of the peripheral nervous system. Maximum ChAT activity was found in the caudate nucleus and the ventral roots of the spinal cord. High activity was also present in the thalamus, the pons, the cerebral cortex, the medulla oblongata, the ventral spinal horns and the sciatic nerve. The lowest activity was measured in the cerebellum, the dorsal cord roots and the spinal ganglia. Maximum AChE activity was found in the caudate nucleus and the cerebellum. Relatively high activity was also present in the thalamus, the pons, the medulla oblongata, the grey matter of the spinal cord and the spinal ganglia. The lowest AChE activity was measured in the ventral and dorsal spinal roots.     

Differential and independent manifestation within co-containing neurones of neuropeptide Y and tyrosine hydroxylase during ontogeny of the rat central nervous system

The development of tyrosine hydroxylase- and neuropeptide Y-immunoreactive cell bodies in the foetal rat brain was analyzed immunohistochemically using antibodies raised against tyrosine hydroxylase and neuropeptide Y. Possible co-existence of these two substance within the same neurones was investigated by comparison of adjacent sections.

In the ventral medulla oblongata, neurones containing both neuropeptide Y- and tyrosine hydroxylase-like immunoreactivity were demonstrable in and around the lateral reticular nucleus as early as the 17th day of gestation. The total number and the proportion of cells exhibiting this co-existence increased from this stage up to birth. In the nucleus of the solitary tract in the dorsal medulla oblongata, NPY-immunoreactive cells bodies were first visualized at day 13 of gestation. However, although tyrosine hydroxylase-immunoreactive cells could also be seen within the nucleus at this and later ages, they occupied a different, more caudal and medial part. Consequently, no neurones containing both neuropeptide Y and tyrosine hydroxylase were apparent up to the day of birth. Finally, in the locus coeruleus, tyrosine hydroxylase-immunoreactive neurones were also demonstrable at day 13 of gestation. In this case, however, no neuropeptide Y-immunoreactive somata could be seen in the nucleus until day 21.

The present study indicates that the existence of neuropeptide Y and tyrosine hydroxylase in co-containing neurones is not inextricably linked, and suggests that the factors controlling the synthesis of these two substances are not identical.     

Ontogeny of central CO2 chemoreception: chemosensitivity in the ventral medulla of developing bullfrogs

Sites of central CO2 chemosensitivity were investigated in isolated brain stems from Rana catesbeiana tadpoles and frogs. Respiratory neurograms were made from cranial nerve (CN) 7 and spinal nerve 2. Superfusion of the brain stem with hypercapnic artificial cerebrospinal fluid elicited increased fictive lung ventilation. The effect of focal perfusion of hypercapnic artificial cerebrospinal fluid on discrete areas of the ventral medulla was assessed. Sites of chemosensitivity, which are active continuously throughout development, were identified adjacent to CN 5 and CN 10 on the ventral surface of the medulla. In early- and middle-stage tadpoles and frogs, unilateral stimulation within either site was sufficient to elicit the hypercapnic response, but simultaneous stimulation within both sites was required in late-stage tadpoles. The chemosensitive sites were individually disrupted by unilateral application of 1 mg/ml protease, and the sensitivity to bath application or focal perfusion of hypercapnia was reassessed. Protease lesions at CN 10 abolished the entire hypercapnic response, but lesions at CN 5 affected only the hypercapnic response originating from the CN 5 site. Neurons within the chemosensitive sites were also destroyed by unilateral application of 1 mM kainic acid, and the sensitivity to bath or focal application of hypercapnia was reassessed. Kainic acid lesions within either site abolished the hypercapnic response. Using a vital dye, we determined that kainic acid destroyed neurons by only within 100 microm of the ventral medullary surface. Thus, regardless of developmental stage, neurons necessary for CO2 sensitivity are located in the ventral medulla adjacent to CN 5 and 10.     

Effects of neuropeptide Y on forskolin, alpha 2- and beta-adrenoceptor-regulated cAMP levels in the rat brain slice.

Neuropeptide Y (10(-6) M) significantly attenuated forskolin-stimulated cAMP levels in slices of the medulla oblongata from WKY rats. No effect of NPY was observed on basal levels of cAMP in this region. Pretreatment with pertussis toxin (2 micrograms and 5 micrograms) IC prevented the reduction of forskolin-stimulated cAMP levels elicited by NPY in the medulla oblongata, suggesting that NPY is acting through an inhibitory guanine nucleotide binding protein to reduce cAMP accumulation. Moxonidine, an alpha 2-adrenoceptor agonist, was observed to reduce forskolin-stimulated cAMP levels in medullary slices. This inhibitory response was attenuated in the presence of NPY (10(-6) M). The beta-adrenoceptor agonist isoprenaline also elevated cAMP levels in the medulla oblongata; however, NPY did not alter this response. It is therefore proposed that the previously reported hemodynamic actions of NPY in the medulla oblongata, an area of cardiovascular significance, may be mediated via a reduction in cAMP levels. Moreover, an interaction between NPY and alpha 2-adrenoceptors, but not beta-adrenoceptors, on cAMP production in the medulla slice preparation was evident.     

Fine structure of the ependyma and intercellular junctions in the area postrema of the rat

Summary Ependymal cells and their junctional complexes in the area postrema of the rat were studied in detail by tracer experiments using horseradish peroxidase (HRP) and colloidal lanthanum and by freeze-etch techniques, in addition to routine electron microscopy. The ependyma of the area postrema is characterized as flattened cells possessing very few cilia, a moderate amount of microvilli, a well-developed Golgi apparatus and rough endoplasmic reticulum. Numerous vesicles or tubular formations with internal dense content were found to accumulate in the basal processes of ependymal cells; the basal process makes contact with the perivascular basal lamina. It is suggested that the dense material in the tubulovesicular formations is synthesized within the ependymal cell and discharged into the perivascular space. The apical junctions between adjacent ependymal cells display very close apposition, with a gap of 2–3 nm, but no fusion of adjacent plasma membranes; they thus represent a transitional form between the zonulae adhaerentes present in the ordinary mural ependyma and the zonulae occludentes in the choroidal epithelium. A direct intercommunication between the ventricular cerebrospinal fluid (CSF) and the blood vascular system indicates that a region exists lacking a blood-ventricular CSF barrier.     

An immunocytochemical and ultrastructural study of a specialized glial region of the medulla oblongata of the adult and juvenile grey mullet

Electron microscopy together with glial fibrillary acidic protein (GFAP) and vimentin immunocytochemistry reveals the presence of a specialized glial region in the octavolateral area of the medulla oblongata of adult and juvenile grey mullets, Chelon labrosus. Glial cells, which can he characterized as ependymal and subependymal cells, originate in the walls of the lateral recess of the fourth ventricle. They contain numerous gliofilaments and are connected through frequent gap junctions. Electron microscopy also reveals the lack of nerve cell parikarya and processes in this region. Immunocytochemistry reveals that the glial cells are strongly GFAP-positive in both adults and juveniles, whereas vimentin is only detected in this region in juveniles. The meninges associated with this glial region contains connective fibres, formed of very thick collagen fibres, that arc metachromatic and PAS-positive under light microscopy. These findings strongly support a structural role for this medullary specialization. Differences between adults and juveniles in the distribution of GFAP- and vimentin-immunoreactive structures in other regions of the medulla oblongata are also reported.     

Age differences in responsiveness of brainstem chemosensitive neurons to extracellular pH changes

The sensitivity of neurons in the caudal chemosensitive area on the ventrolateral surface of the medulla oblongata (VMS) to extracellular pH changes was examined in newborn and young developing kittens and compared to that of adult cats. The pH was varied by superfusion of the VMS with mock cerebrospinal fluid (CSF) of pH 7.4 (control), 7.0 (acid) and 7.8 (alkaline). A total of 97 neuronal units in the three age groups changed their firing rates inversely in response to extracellular fluid (ECF) pH changes. The greatest sensitivity was found in the adult group where acid superfusion caused an increase in neuronal activity. The least sensitivity was observed in the newborn group (1-6 days old), whereas the young kitten group (4-6 weeks old) exhibited an intermediate sensitivity. Neurons of kittens older than 7 weeks of age demonstrated a response pattern characteristic of the adult group. Neurons of neonates older than seven days, exhibited a response pattern characteristic of the young kitten group.