Adult Rat CNS Research Articles

Is there capacity for anatomical and functional repair in the adult somatosensory thalamus?

The capacity for structural and functional remodeling in damaged adult CNS sensory systems can be studied by replacement of neurons in damaged structures by fetal cells from these anatomical origins. For integration to take place, the replacement paradigm assumes that (a) reconnection of adult host afferent fibers onto developing neurons is possible and (b) that the correct molecular signals exist also in the adult brain for fetal neurons to extend axons and pattern synaptic contacts. We have tried to answer some of these fundamental questions by using neuronal depletion models followed by neuronal replacement in the adult rat CNS (Isacson et al. 1984. Nature (London) 311: 458–460; Isacson et al. 1988. Prog. Brain Res. 78: 13–27; Nothias et al. 1988. Brain Res. 461: 349–354; Peschanski and Isacson. 1988. J. Comp. Neurol. 274: 449–463; Sofroniew et al. 1990. Prog. Brain Res. 82: 313–320). In one such model, kainic acid infusions deplete the ventrobasal complex (VB) of all neurons projecting to the somatosensory cortex, while afferent axons from the lemniscal and monoaminergic systems remain in the area. Direct implantation of fetal neurons (gestation age 15–16) of ventrobasal destination allows reconnection of circuitry to take place at the thalamic level, as studied by anatomical tracers, electron microscopy, and functional 2-deoxyglucose studies, while fetal thalamic VB neurons appear less likely to growt through the internal capsule toward the cortical level.

Maturation and fine structure of thalamic reticular neurons transplanted into the adult rat CNS.

This work was undertaken to determine whether or not cell suspension transplants provide a promising vehicle for the in vivo study of neuronal development. Cell suspensions of rat fetal tissue that contained both reticular (RT) and ventrobasal primordia were transplanted into the excitotoxically lesioned somatosensory thalamus of adult rats. The fine structure of transplanted GABAergic RT neurons that were identified by immunocytochemistry was investigated 5 days to 7 months following transplantation. The dissociation and transplantation of the tissue results in disruption of the extracellular matrix and alteration of cellular interactions which could introduce major changes in neuronal maturation. Our results show that maturation of transplanted GABAergic RT neurons was comparable to normal and that they were able to establish and receive, with one exception, usual sets of connections. In contrast, disruption of the tissue was responsible for the definitive loss of organotypic characteristics and the emergence of abnormal connections, especially synaptic connections from somatosensory afferents, the functional significance of which will be important to determine.

Ciliary neurotrophic factor prevents neuronal degeneration and promotes low affinity NGF receptor expression in the adult rat CNS

Recombinant human ciliary neurotrophic factor (CNTF) was infused for 2 weeks into the lateral ventricle of fimbria-fornix transected adult rats, and its effects were compared with those of purified mouse nerve growth factor (NGF). We provide evidence that CNTF can prevent degeneration and atrophy of almost all injured medial septum neurons (whereas NGF protects only the cholinergic ones). CNTF is also involved in up-regulation of immunostainable low affinity NGF receptor (LNGFR) in cholinergic medial septum and neostriatal neurons and in a population of lateral septum neurons. In contrast to NGF, CNTF did not stimulate choline acetyltransferase in the lesioned septum and normal neostriatum (pointing to different mechanisms for the regulation of choline acetyltransferase and LNGFR), cause hypertrophy of septa) or neostriatal cholinergic neurons, or cause sprouting of LNGFR-positive (cholinergic) septal fibers.

Developmental and Age‐Dependent Changes of 28‐kDa Calbindin‐D in the Central Nervous Tissue Determined with a Sensitive Immunoassay Method

For the quantitative analysis of vitamin D-dependent 28-kDa calcium-binding protein (calbindin-D) in the CNS, we have established a highly sensitive immunoassay method. The antisera were raised in rabbits with purified calbindin-D from rat kidneys, and the antibodies were purified with a calbindin-D-coupled Sepharose column. The purified antibodies were specific for calbindin-D, showing a single band on the immunoblot with the extract of rat kidney or cerebellum. The sandwich-type immunoassay system was prepared by the use of purified monospecific antibodies, and the minimum detection limit of the assay was 0.1 pg or 3.6 amol of calbindin-D, which was sufficiently sensitive for the measurement of calbindin-D content in isolated Purkinje cell bodies at the level of single cells. The average content of calbindin-D in a single Purkinje cell was 0.05 pg. Calbindin-D was detected in most of the rat tissues examined, but it was present predominantly in the kidney and CNS, especially in the cerebellum. Calbindin-D was detected at a similarly low level in the cerebral cortex, cerebellum, and brainstem of rat embryos of 15 gestational days, and it increased gradually but differently in these regions, reaching the respective adult levels by 4-5 weeks of postnatal age. In contrast, kidney calbindin-D increased sharply between 15 gestational days and 3 postnatal days, reaching the adult level by 6 days of age. Calbindin-D levels in the adult rat CNS were affected little by age, whereas the concentrations in human cerebral cortices were significantly low in the aged brain as compared with those in the young brain.(ABSTRACT TRUNCATED AT 250 WORDS)

Reactive astrocytes are substrates for the growth of adult CNS axons in the presence of elevated levels of nerve growth factor

To assess the necessary parameters for the growth of axons within the adult rat CNS, we have used intracerebral grafts of primary fibroblasts genetically engineered to express nerve growth factor (NGF). Following the implantation of NGF-producing primary fibroblasts within the striatum, cholinergic axons arising from the nucleus basal is grow toward and penetrate these grafts between 1 and 8 weeks. Grafts of noninfected control cells do not elicit axon sprouting at any time. Unmyelinated axons grow into grafts of NGF-producing cells only on reactive astrocytic processes, which contribute to a surrounding glial border. From our data concerning axon growth within the adult rat CNS, we conclude that reactive astrocytes can act as conducive substrates for growing axons; and only in the presence of elevated levels of NGF will permissive substrates (e.g., astrocytes) support axon growth by NGF-sensitive neurons.

Choline acetyltransferase messenger RNA expression in developing and adult rat brain: regulation by nerve growth factor

The polymerase chain reaction (PCR) was used to develop a method for detection and relative quantification of the choline acetyltransferase (ChAT) mRNA in neonatal and adult rat CNS. Oligonucleotide primers derived from a porcine ChAT cDNA sequence were used in coupled reverse transcriptase (RT)-PCR to amplify a cDNA sequence of 206 bp which arises in a cycle- and RNA-dependent manner and which hybridizes with both an internal oligonucleotide and a ChAT cDNA probe. ChAT mRNA was detected in spinal cord, septal area, striatum, cortex and hippocampus but not in cerebellum and cardiac or skeletal muscle. In the septal area, relative quantitative evaluation of ChAT mRNA levels by RT-PCR indicates that this transcript is developmentally regulated and increased following intracerebral administration of nerve growth factor (NGF) to both neonatal and young adult rats. This suggests that the increases of ChAT activity observed in basal forebrain during development or after NGF administration are, at least in part, associated with an increase in corresponding levels of mRNA.

C-JUN-like immunoreactivity in the CNS of the adult rat: Basal and transynaptically induced expression of an immediate-early gene

An immunocytochemical study of dorsal root ganglia, spinal cord and medulla oblongata was performed with antisera against the c-jun proto-oncogene encoded protein. The c-JUN-like immunoreactivity was restricted to the cell nucleus. In the CNS of untreated rats a basal c-JUN-like immunoreactivity was present in the nuclei of two types of neurons: motor and autonomic. Labelled nuclei could be seen in many motoneurons of the ventral horn of the entire length of spinal cord and the lower medulla oblongata, as well as in the area of the nucleus hypoglossus, the dorsal motor nucleus of nucleus vagus, nucleus ambiguus, nucleus facialis, nucleus abducens and motor nucleus of nucleus trigeminus. Additionally, labelled nuclei were found in the preganglionic sympathetic and preganglionic parasympathetic cells of the nucleus intermediolateralis and nucleus intercalatus in the spinal cord. In the medulla oblongata we found a cluster of cells with c-JUN-like immunoreactivity in an area between the dorsomedial part of the oral nucleus spinalis trigeminalis and the lateral border of the knee of facial nerve. Additionally, a second cluster of c-JUN-like immunoreactivity cells was visible between the ventromedial part of the oral nucleus spinalis trigeminalis and the lateral border of the rostral nucleus facialis. Examination of the characteristics of all cell groups with a basal c-JUN-like immunoreactivity in the spinal cord and lower brainstem revealed an overlapping distribution with cholinergic cell groups. Basal c-JUN-like immunoreactivity was also seen in the dorsal root ganglion cells. We examined the factors which can effect the expression of the c-JUN protein. Maximal expression of c-JUN-like immunoreactivity was observed after electrical stimulation of primary afferents. Stimulation of sciatic nerve at a strength sufficient to recruit Aδ- and C-fibres produced c-JUN-like immunoreactivity in many nuclei of the ipsilateral dorsal horn of the lumbar spinal cord. c-JUN-like immunoreactivity was first detectable at 30 min following the end of stimulation, reached a maximum after 1 h, remained unchanged for another l h and declined to the basal level after 16h. The distribution of c-JUN-like immunoreactivity in the lumbar cord coincided with the region of termination of sciatic nociceptive afferents. Contralateral c-JUN-like immunoreactivity appeared after 4h. After noxious mechanical stimulation of the plantar hindpaw c-JUN-like immunoreactivity occurred in the spinal area of termination of nociceptive afferents of the tibial nerve. Noxious stimulation did not provoke additional c-JUN-like immunoreactivity in dorsal root ganglia. Electrical stimulation of large myelinated Aα/β-fibres of sciatic nerve and non-noxious stimulation of peripheral sensory nerve receptors was not effective for induction of c-JUN-like immunoreactivity in spinal neurons. This is the first study demonstrating the expression c-JUN protein in vivo, a transcriptional operating protein with a functional relation to the extensively studied c-FOS protein. Our data demonstrate a basal constitutive expression of c-JUN in CNS of normal adult rat which is not paralleled by the expression of c-FOS protein, thus indicating a fundamental selective expression of these functionally related proteins. Furthermore, we reveal for the first time that transient expression of immediate early genes in spinal cord is a neurogenic consequence during nociception which is dependent upon the activation of nociceptive primary afferents.

Glial changes following an excitotoxic lesion in the CNS—I. Microglia/macrophages

When an area of the adult rat CNS is depleted of neurons by an in situ excitotoxic injection, afferent axons to the area exhibit morphological alterations reminiscent of growth cones. These morphological changes are likely to be related to the deprivation of target cells. In addition, however, the area of neuronal loss is itself the site of profound changes in glial cell content, and altered axon-glial interactions may play a role in the axonal changes. In an attempt to define these interactions, we have undertaken a systemic study of glial populations in excitotoxically lesioned CNS over time. The microglial/macrophagic response is analysed in this paper; the astrocytic response is described in the companion paper [Dusart et al. (1991) Neuroscience 45, 541–549]. The microglial/macrophagic response was studied following kainic acid-induced neuronal loss in the thalamus of the adult rat. These microglial/macrophagic cells were labeled with the B4 isolectin from Griffonia simplicifolia, and the time-course of their response was studied between one day and one year post-lesion. This time-course study revealed different stages in the evolution of the response. At one day post-lesion, cell counts indicated that there was no increase in the number of non-neuronal cells in the neuron-depleted area. However, activated labeled cells were present in the entire thalamus on the side of the lesion, neuron-depleted or not. They were characterized by both increased lectin-binding and altered morphology when compared to quiescent microglia. In the absence of recruitment and/or proliferation, this result indicates that the early response consisted solely of the activation of resident microglia. By contrast, we observed a progressive increase in the number of non-neuronal cells in the lesion from four to 15 days post-lesion. A recruitment of blood-borne monocytes was apparent, and the observation of mitotic labeled cells indicated a proliferation of microglial/macrophagic cells in situ. There was a progressive decrease in the microglial/macrophagic reaction that began one month after lesion. In a thin band of parenchyma surrounding the neuron-depleted area, activated microglial/macrophagic cells were seen contacting neurons, and clusters of glial cells were observed around neurons up to one year post-lesion. These results suggest that neurons around the lesion site itself may be injured, secondarily, from a long term deleterious effect of the inflammatory process. This study allows us to conclude that activated microglia/macrophages are the predominant glial cell type in the excitotoxically lesioned CNS over the first weeks. It is known that, in their activated stage, these cells are able to release numerous factors acting on other cell types. It is proposed, therefore, that microglia/macrophages may play a key role in the evolution of cellular populations observed in the area of neuronal loss.

Localization of FGF receptor mRNA in the adult rat central nervous system by in situ hybridization

Fibroblast growth factor receptor (FGF-R) mRNA expression was examined in the adult rat CNS. Northern blot analysis showed a distinct 4.3 kb transcript in various CNS regions. In situ hybridization revealed widely distributed, but specific, populations of cells that express FGF-R mRNA. The most intense hybridization signals were observed in the hippocampus and in the pontine cholinergic neurons. The limbic system and brainstem nuclei, including motor nuclei, showed robust labeling. Cerebellar granule cells and spinal cord neurons were positive for FGF-R mRNA. The distribution of FGF-R mRNA differed significantly from that of NGF receptor mRNA; particularly, no hybridization signal was detected in basal forebrain cholinergic neurons. These results strongly suggest that FGF or FGF-like molecules may exert effects on specific neuronal populations in the mature CNS.

Nerve growth factor infusion into the denervated adult rat hippocampal formation promotes its cholinergic reinnervation

The well-documented but little-understood failure of lengthy axonal regeneration after injury of the adult mammalian CNS may be caused by an insufficient availability of local growth-promoting factors. If so, identifying and supplying the missing factors may result in better central axonal regeneration. This hypothesis was tested in an adult rat CNS model in which peripheral nerve grafts were placed into a lesion cavity between the septum and hippocampal formation. Continuous infusion of nerve growth factor (NGF) into the dorsal hippocampal tissue dramatically enhanced and accelerated the regrowth and penetration of cholinergic axons into the hippocampal formation. Thus, NGF can overcome the apparent resistance of the hippocampal CNS tissue to cholinergic reinnervation.

Isolation, culture, and characterization of adult rat oligodendrocytes

Investigation into CNS demyelinating diseases, which usually occur in adults, can be facilitated by the use of a good in vitro model. We have established a methodology whereby oligodendrocytes from adult rat CNS can be cultured in vitro, and we have characterized these cultures morphologically and immunologically. Approximately 1 g of spinal cord and brainstem per adult rat was removed and dissociated mechanically and enzymatically. After filtration of the white matter homogenate, myelin was removed by 0.9 M sucrose density centrifugation. The cells were further purified by centrifugation through a 0.3%/4% discontinuous gradient of bovine serum albumin (BSA). The pellet was resuspended and placed in an untreated 6-well culture dish overnight to allow the astrocytes to attach. The non-adherent cells were replated on poly-l-lysine-treated coverslips. Approximately 8.25 x 10(5) cells were recovered per animal. The adult oligodendrocytes initially appeared as rounded cell bodies, but after 2-5 days in vitro (DIV), the oligodendrocytes extended 6-10 thick processes. A membrane sheath between these processes was immunostainable with either anti-galactocerebroside (GC), anti-O4, anti-myelin basic protein (MBP), or anti-2'3' cyclic nucleotide 3' phosphohydrolase (CNPase) and was also evident in scanning EM. Older cultures (up to 60 DIV) maintained whorls of myelin and transmission EM revealed a major dense line distance of approximately 103 A with up to 11 concentric layers of membrane. Immunologically, the adult oligodendrocytes are GC+, O4+, MBP+, CNPase+, and GFAP-. The method described will allow adult rat oligodendrocytes to be isolated and maintained in culture; these cultures retain the characteristics of differentiated adult oligodendrocytes.

Laminin-like antigen in rat CNS neurons: Distribution and changes upon brain injury and nerve growth factor treatment

Using several antibodies against rat or human laminin and an avidin-biotin immunocytochemical protocol, laminin-like immunoreactivity was detectable in the rat nervous system in expected locations, i.e., associated with blood vessels and reactive astrocytes. However, laminin staining was also abundantly present within neuronal cell bodies in most parts of the developing and adult rat CNS. Medial septum neuronal immunoreactivity was lost after septo-hippocampal disconnection, but could be preserved or even restored by intraventricular administration of nerve growth factor. Thus, at least for medial septum neurons, this laminin-like molecule can be accumulated or produced independent of direct hippocampal (target) contact. It remains to be determined whether CNS neuronal “laminin” pocesses activities similar to those found for laminin in vitro.

Differential Ultrastructural Localization of Myelin Basic Protein, Myelin/Oligodendroglial Glycoprotein, and 2′,3′‐Cyclic Nucleotide 3′‐Phosphodiesterase in the CNS of Adult Rats

In a light and electron microscopic immunocytochemical study we have examined the distribution of myelin basic protein (MBP), 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP), and myelin/oligodendroglial glycoprotein (MOG) within CNS myelin sheaths and oligodendrocytes of adult Sprague-Dawley rats. Ultrastructural immunocytochemistry allowed quantitative analysis of antigen density in different myelin and oligodendrocyte zones: MBP was detectable in high density over the whole myelin sheath, but not in regions of loops, somata, or the oligodendrocyte plasma membrane. CNP reactivity was highest at the myelin/axon interface, and found in lower concentration over the outer lamellae of myelin sheaths, at the cytoplasmic face of oligodendrocyte membranes, and throughout the compact myelin. MOG was preferentially detected at the extracellular surface of myelin sheaths and oligodendrocytes and in only low amounts in the lamellae of compacted myelin and the myelin/axon border zone. Our studies, thus, indicate further the presence of different molecular domains in compact myelin, which may be functionally relevant for the integrity and maintenance of the myelin sheath.

The monoclonal antibodies Elec-39, HNK-1 and NC-1 recognize common structures in the nervous system and muscles of vertebrates

The IgM monoclonal antibodies, Elec-39, HNK-1 and NC-1, recognize the same subset of Torpedo electric organ acetylcholinesterase (AChE). We show that they react against a glycosphingolipid (SGPG) containing a sulfated glucuronic acid (SGA). The three antibodies appear essentially identical in their specificity but differ in their affinity for the antigens. We have examined their binding in the CNS, nerves and muscles of several vertebrate species, at the optical and in some cases at the electron microscope level. All three antibodies label the same structures: they show diffuse staining around neuromuscular endplates and label the plasma membrane of the Schwann cells, surrounding the outer layer of myelin sheaths. In the adult rat CNS, the antibodies label certain defined structures, notably extracellular material in the habenula and in the CA2 layer of the hippocampus. In the cortex and cerebellum, they label the surface of neural processes and terminals apposed to large multipolar neurons and Purkinje cells, as well as membranous material contained in inclusions dispersed in the cytoplasm of these neurons. These localizations are consistent with the suggestion that the SGA-antigens may be involved in cellular interactions.

Rapid growth of host afferents into fetal thalamic transplants

Fetal cell suspension grafts grow and differentiate when implanted into adult rat CNS areas previously neuron-depleted using an excitotoxin. There is some controversy in the literature concerning the timetable of establishment and possible extent of host-graft connections in these experimental conditions. The present study was undertaken to analyze the development of adult host monoaminergic afferents into a transplant formed by fetal thalamic neurons in the previously excitotoxically lesioned thalamus. It is demonstrated that both norepinephrin- and serotonin-immunoreactive fibers are present in the transplant as soon as 8 days after grafting. At those times, immunoreactive fibers exhibit morphological characteristics typically associated with immature stages. After longer survival time, up to 4 weeks after grafting, immunoreactive fibers are numerous in the transplant and exhibit morphological features comparable to those observed in the adult thalamus. These results demonstrate the rapid ingrowth of some fiber systems of the adult host into the transplant and suggest that grafted fetal cells can be functionally integrated into the host circuitry as soon as a few weeks after grafting.

Nerve fiber growth in culture on tissue substrata from central and peripheral nervous systems

In adult mammals, injured neurons regenerate extensively within the PNS but poorly, if at all, within the CNS. We have studied the effect of substrata consisting of tissue sections from various nervous systems on nerve fiber growth in culture and correlated our results with the growth potential of these tissues in vivo. Ganglionic explants from embryonic chicks (9-12 d) fail to extend nerve fibers onto sections of adult rat optic nerve or spinal cord (CNS) but do so on sciatic nerve (PNS). Dissociated DRG neurons behave similarly whether in serum-containing or defined medium. Tissue substrata from nervous systems that support regeneration in vivo--i.e., goldfish optic nerve, embryonic rat spinal cord, degenerating sciatic nerve--also support fiber growth in culture. Within the same culture, neurons will grow onto sciatic nerve rather than neighboring optic nerve sections, suggesting that the responsible agent(s) is not soluble. In addition, neurons adhere more extensively to sciatic nerve substrata than to optic nerve. The occurrence of 3 molecules known to be involved in neuron-substratum adhesion and nerve fiber growth in vitro has been documented immunocytochemically in the tissue sections. One of these, laminin, is demonstrable in all tissues tested that supported nerve fiber growth. Immunoreactivities for fibronectin and heparan sulfate proteoglycan are found in only some of these tissues. None of these 3 molecules can be demonstrated in neural cells of normal adult rat CNS tissue. Our data suggest that these molecules may be important effectors of nerve regeneration in neural tissues.

G D3 ganglioside is a glycolipid characteristic of immature neuroectodermal cells

Biochemical studies have indicated that the disialoganglioside, GD3, is a major glycolipid component of the immature vertebrate CNS, but a minor element within the mature CNS. We have investigated its cellular localization in rat CNS by immunofluorescence using a mouse monoclonal antibody that recognizes GD3. In tissue sections of postnatal CNS, the antibody bound to cells in several areas known to contain immature neuroectodermal populations: the subventricular zone beneath the lateral ventricle, the external germinative layer of the cerebellar cortex, and the dentate gyrus of the hippocampus. GD3-containing cells were also found in developing white matter of the forebrain and cerebellar folia. Using a double-label immunofluorescence method, we found that the GD3-positive white matter cells did not express the astrocytic marker, glial fibrillary acidic protein. In adult rat CNS, we could not detect antibody binding to neurons or glia. Scattered GD3-positive cells were apparent in the adult subventricular zone. Our results indicate that GD3 ganglioside is a membrane component characteristically expressed in the rat CNS by neuroectodermal stem cells, both neuronal and glial precursors.

Specific and potent mitogenic effect of axolemmal fraction on schwann cells from rat sciatic nerves in serum-containing and defined media

Using cultures of Schwann cells from neonatal rat sciatic nerves, we examined the mitogenic activity of an axolemmal fraction from adult rat CNS. Axolemmal fraction proved a potent mitogen, stimulating [ 3H]thymidine incorporation into Schwann cell DNA 13.5 fold over control values when axolemmal fraction equivalent to 16 μg of protein per culture microwell or more was added. Half maximal stimulation was obtained with addition of axolemmal fraction equivalent to 4 μg of protein. The concentration-dependence and magnitude of the mitogenic response of the cultured cells were nearly identical whether they were maintained in vitro for 1 day or for 2 weeks prior to addition of the axolemmal fraction. A study of the time-course of the effect of axolemmal fraction on Schwann cell mitosis showed that maximal [ 3H]thymidine incorporation took place during the fifth day after addition of axolemmal fraction. Axolemmal fraction also produced stimulation of [ 3H]thymidine incorporation into Schwann cells, seeded and cultured in a serum-free defined medium. Though the concentration-dependence of the mitogenic effect in the absence of serum was similar to that in a serum-containing medium, maximal stimulation in the defined medium was only 2.8-fold. The mitogenic activity of axolemmal fraction was rapidly and almost totally inactivated by sonication or homogenization, and was partially lost after exposure to heat. The mitogenic activities of plasma membrane fragments from rat skeletal muscle or rat erythrocytes, andof mitochondrial fragments (the major contaminant of the axolemmal fraction) were one-tenth that of axolemmal fraction or less. In contrast to glial growth factor prepared from bovine pituitaries (GGF-BP), which stimulates proliferation of both fibroblasts and Schwann cells, axolemmal fraction induced proliferation of Schwann cells but not of endoneurial fibroblasts; cultures treated with axolemmal fraction demonstrated a 3-fold increase in Schwann cell population in 10 days without detectable increase in number of fibroblasts. Also in contrast to GGF-BP, the mitogenic effect of which is considerably enhanced by simultaneous addition of cholera toxin to the medium, cholera toxin had no effect on the Schwann cell proliferative response to axolemmal fraction.

Insulin- and glucagonlike peptides in the brain.

The cellular localization and regional distribution of insulin- and glucagonlike substance, C-peptide-like immunoreactivity, thiol:protein disulphide oxidoreductase, TPO (E.C.1.8.4.2.), and insulin/glucagon-specific proteinase, ISP (E.C.3.4.22.-), are studied in the CNS of man, adult and juvenile rats, mice, tortoises, and frogs by use of immunohistochemistry. Furthermore, the content of immunoreactive insulin, glucagon, and C-peptide was estimated in human cadaver brains by radioimmunoassay. It could be shown that insulinlike immunoreactive material is widely distributed in the human brain and the CNS of juvenile rats as well as in mice, whereas in the CNS of adult rats and nonmammalian animals (frogs, tortoises) the polypeptide is restricted to a few nerve cell populations. C-peptide immunoreactivity was demonstrated in human CNS in the same nerve cells as insulin. By use of two different glucagon-antisera it was revealed that gut-type glucagon occurs in many nerve cells of human and mouse brains, as well as in the CNS of juvenile rats. On the other hand, pancreas-type glucagon was less widely distributed in the human brain and nearly not detectable in the CNS of mice and rats. With the exception of neurosecretory nerve cells, there was a high degree of coincidence between the localization of insulin and TPO. The immunoreaction against the ISP antiserum was weak, but correlated well with the distribution of insulin-immunoreactivity. The occurrence of TPO and ISP in the brain demonstrates the ability of nervous tissue to degrade insulin and glucagon. By radioimmunoassay it was established that human brain contains insulin, glucagon and C-peptide at concentrations that exceed blood levels. We conclude from our data that, at least in part, cerebral insulin and glucagon are products of the brain itself.

Influence of the membrane fluidity on the adenosine receptor sites, cAMP system coupled, in CNS of adult and newborn rats

Influence of the membrane fluidity on the adenosine receptor sites, cAMP system coupled, in CNS of adult and newborn rats