Biomedical subjects

H Steinbusch

Publications and source records attributed to H Steinbusch.

At least 19 recordsLinked to original sources

PubMed2004

Dual effect of diazepam on cGMP levels in rat brain slices.

The effect of diazepam on NO-mediated cGMP synthesis was studied in rat brain slices. It was found that diazepam dose-dependently decreased cGMP synthesis in cerebellar slices, with an inhibition of 90% at 1 mM diazepam. cGMP levels in the presence of diazepam were not restored to control levels by the addition of 0.1 mM sodium nitroprusside, whereas the decrease in cerebellar cGMP levels induced by 0.1 mM L-NAME was restored by the simultaneous application of NO-donors. In addition to the decrease of cGMP levels in neuronal structures induced by 1 mM diazepam, we observed increased cGMP immunoreactivity in glial cells in the cerebellum, the hippocampus, and the cerebral cortex. The significance of this observation is discussed.

1-Methyl-3-isobutylxanthine↗

PubMed2003

Decreased levels of cGMP in vitreous and subretinal fluid from eyes with retinal detachment.

BACKGROUND: Cyclic guanosine monophosphate (cGMP) is produced in different retinal cells, including photoreceptor cells, wherein cGMP mediates photo-transduction. CGMP is degraded by phosphodiesterases (PDE). The aim was to investigate whether retinal detachment alters intraocular cGMP levels in human eyes. METHODS: cGMP and PDE were determined in vitreous fluid from 50 eyes with a retinal detachment (group I) and in 20 control samples (group II) of vitreous fluid from eyes without retinal detachment. Group III consisted of subretinal fluid samples from 70 eyes with retinal detachment. RESULTS: cGMP in vitreous fluid from eyes with retinal detachment (6.5 (SD 1.7) nM) was decreased compared to controls (67.1 (10.0) nM) (p<0.0001). In subretinal fluid, the mean level of cGMP was 2.4 (0.2) nM. No PDE could be detected in any of the intraocular fluid samples of patients nor controls. A decrease in the mean level of cGMP in subretinal fluid of eyes with retinal detachment correlated with a longer duration of detachment (r = -0.45, p = 0.007). CONCLUSIONS: Retinal detachment was found to be associated with a decrease in vitreous cGMP concentration. In subretinal fluid, a low cGMP level correlated inversely with the duration of the detachment.

Adolescent↗

PubMed2001

Presence of soluble and particulate guanylyl cyclase in the same hippocampal astrocytes.

The localisation of particulate and soluble guanylyl cyclase was studied in hippocampal astrocytes. Counting the colocalisation of cGMP immunoreactivity with the astrocytic marker glial fibrillary acidic protein after stimulation of brain slices with sodium nitroprusside (0.1 mM) or atrial natriuretic peptide (100 nM), we were able to show that at least 67% of the hippocampal astrocytes contained both guanylyl cyclase isoforms. In addition, it was shown that a large number of atrial natriuretic peptide, brain-derived natriuretic peptide or sodium nitroprusside responsive cells contain the beta1-subunit of the soluble guanylyl cyclase. The results show that, in at least a subset of hippocampal astrocytes, soluble and particulate guanylyl cyclases are simultaneously present in the same cells.

1-Methyl-3-isobutylxanthine↗

PubMed2001

Levels of basic fibroblast growth factor, glutamine synthetase, and interleukin-6 in subretinal fluid from patients with retinal detachment.

PURPOSE: To investigate the presence of basic fibroblast growth factor, glutamine synthetase activity, and interleukin-6 in subretinal fluid from patients with retinal detachment. METHODS: In a prospective study we measured basic fibroblast growth factor, glutamine synthetase activity, interleukin-6, and total protein in subretinal fluid samples from 96 eyes from 94 consecutive patients with a retinal detachment corrected by a conventional scleral buckling operation in our clinical practice. As controls, vitreous fluid samples from eyes with a macular hole (n = 6) or pucker (n = 11) were used. Laboratory data of the patient group were compared with the control group and correlated with various clinical data. RESULTS: Levels (median, range) of basic fibroblast growth factor, glutamine synthetase activity, interleukin-6, and total protein were significantly higher in patients than in controls (P <.0001). An increased level of glutamine synthetase and total protein correlated with a longer duration of the retinal detachment (r =.4, P =.002, and r =.34, P =.001, respectively). Interleukin-6 and basic fibroblast growth factor levels did not correlate with the duration of the detachment. After multivariate logistic regression analysis, no significant relation was found between any of the tested subretinal proteins and a low visual outcome or redetachment. CONCLUSIONS: We found increased levels of basic fibroblast growth factor and glutamine synthetase in subretinal fluid from patients with retinal detachment. Basic fibroblast growth factor and glutamine synthetase may play a role in the pathogenesis and recovery after retinal detachment. The questions of whether the increased levels of basic fibroblast growth factor and glutamine synthetase result from leakage of dying glia cells (including Müller cells) and neurons and if basic fibroblast growth factor is actively produced to protect the photoreceptor cells need further research.

Adolescent↗

PubMed2000

Localization and age-related changes of nitric oxide- and ANP-mediated cyclic-GMP synthesis in rat cervical spinal cord: an immunocytochemical study.

An immunocytochemical technique was used to study the localization and developmental aspects of cyclic GMP (cGMP)-synthesizing structures in the cervical spinal cord of 2-week and 3-month-old Lewis rats in response to the nitric oxide (NO) donor sodium nitroprusside (SNP) and/or atrial natriuretic peptide (ANP). By using cell-specific markers, the cell structures involved were investigated. To visualize cGMP, a combined technique of low- and high-power magnification, using a confocal laser scanning microscope was used. NOS-mediated cGMP synthesis was observed in the cervical spinal cord in laminae I, II and III in 14-day-old rats, which activity was mainly absent at the age of 3 months. The involvement of NO in the NMDA-mediated increase in cGMP immunostaining (cGMP-IS) was demonstrated by the absence of cGMP-IS in slices incubated in the presence of NMDA together with the NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME). This NO-mediated effect of NMDA on cGMP-IS was completely absent in the 3-month-old rats. ANP-mediated cGMP synthesis resulted in an increase in cGMP in laminae I and II, which was generally similar at both ages. Astrocytes in both white and gray matter were found to be cGMP-IS in the basal, NO- and ANP-stimulated conditions. Using confocal laser microscopy, NO-mediated cGMP synthesis was observed in large cholinergic terminals nearby motor neurons in the ventral horn. An extensive colocalization between NO-stimulated cGMP synthesis and parvalbumin-positive (GABAergic) neurons and fibers was observed in all laminae. In the ANP-stimulated condition, a colocalization with parvalbumin structures was found in laminae II and III. No NO- or ANP-mediated cGMP synthesis was found in fibers immunopositive for the presynaptic glutamate transporter, serotonin, or tyrosine hydroxylase.

Age Factors↗

PubMed2000

Nitric oxide synthase does not mediate neurotoxicity after an i.c.v. injection of streptozotocin in the rat.

In the present study we evaluated the possible role of nitric oxide (NO) in mediating neuronal damage in middle-aged rats after an i.c.v. injection of streptozotocin (STREP). An i.c.v. injection of STREP has been reported to decrease the central metabolism of glucose. This inhibition of the energy metabolism after STREP treatment might induce an excitotoxic mechanism, which may lead to the stimulation of NO synthase and, consequently to the synthesis of NO. On the other hand, STREP might induce oxidative stress directly by liberation of NO from its nitroso moiety. To investigate whether NO synthase is involved in a possible excitotoxic mechanism after STREP treatment, some of the rats treated with STREP (1.25 mg/ kg in 4 microl, bilaterally 2 microl/injection site) were also treated with the NO synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME, 20 mg/kg i.p. 10 min, 6, 24 and 96 h after STREP injection). To investigate whether NO liberated from STREP may be responsible for neurotoxic effects, one additional group of control rats received an i.c.v. injection of the NO donor sodium nitroprusside (SNP, 10 microg in 4 microl). We found that STREP affected the behavioral performances in the open field and two-way active avoidance task. In addition, immunostaining for glial fibrillary acidic protein, an indicator of reactive astroglial changes to neuronal damage, showed that this was mainly located in peri- and paraventricular regions of the third and lateral ventricles, like for instance in the septum, caudate putamen and hippocampus. L-NAME treatment had no protective effect on the behavioral impairments and neuronal damage of STREP-treated rats. This suggests that the neuronal damage of STREP may still be a result of the decrease in the central energy metabolism, but without the involvement of NO synthase. This was supported by measuring, using immunostaining, the NO-mediated cyclic GMP production by the enzyme soluble guanylyl cyclase in cortical slices, i.e. L-NAME did not prevent NO production after STREP administration in vitro. In addition, it was found that SNP liberated NO in vitro, whereas in vivo SNP administration did not lead to any behavioral and neuronal deficits at all. However, the present study cannot exclude the involvement of NO liberated from STREP in neuronal damage.

Animals↗

PubMed1999

Down-regulation of ACTH and glucocorticoid receptor immunoreactivity in hypothalamic arcuate neurons after adrenalectomy in the rat.

The expression of glucocorticoid receptor (GR) in rat adrenocorticotropin (ACTH)-containing neurons in rat brain was immunohistochemically investigated. ACTH-containing cell bodies were found mainly in the arcuate nucleus. Most of these neurons exhibited GR immunoreactivities in their nuclei. ACTH-containing nerve fibers were distributed in the bed nucleus of the stria terminalis, periventricular nucleus, retrochiasmatic nucleus, parvocellular part of paraventricular nucleus and dorsomedial hypothalamic nucleus. After adrenalectomy there was a marked decrease of ACTH immunoreactivity, as well as GR immunoreactivity, in neurons of the arcuate nucleus, but ACTH immunoreactivity in the fibers was not affected. These results indicate that glucocorticoids up-regulate ACTH and GR production in hypothalamic arcuate neurons, but that glucocorticoid-induced changes could be delayed in the fibers derived from these neurons.

Adrenal Glands↗

PubMed1999

Local inhibition of hippocampal nitric oxide synthase does not impair place learning in the Morris water escape task in rats.

Recent studies have provided evidence that nitric oxide (NO) has a role in certain forms of memory formation. Spatial learning is one of the cognitive abilities that has been found to be impaired after systemic administration of an NO-synthase inhibitor. As the hippocampus has a pivotal role in spatial orientation, the present study examined the role of hippocampal NO in spatial learning and reversal learning in a Morris task in adult rats. It was found that N omega-nitro-L-arginine infusions into the dorsal hippocampus affected the manner in which the rats were searching the submerged platform during training, but did not affect the efficiency to find the spatial location of the escape platform. Hippocampal NO-synthase inhibition did not affect the learning of a new platform position in the same water tank (i.e. reversal learning). Moreover, no treatment effects were observed in the probe trials (i.e. after acquisition and after reversal learning), indicating that the rats treated with N omega-nitro-L-arginine had learned the spatial location of the platform. These findings were obtained under conditions where the NO synthesis in the dorsal hippocampus was completely inhibited. On the basis of the present data it was concluded that hippocampal NO is not critically involved in place learning in rats.

Animals↗

PubMed1998

The NO-cGMP pathway in the rat locus coeruleus: electrophysiological, immunohistochemical and in situ hybridization studies.

The effect of two nitric oxide (NO) donors, SIN-1 and DEA/NO, as well as of the inactive SIN-1 derivative molsidomin, was studied on locus coeruleus (LC) neurons in a slice preparation using intracellular recordings. In addition, the effect of the guanylate cyclase inhibitor ODQ was analysed. Furthermore, the effect of NO donors on cyclic guanosine monophosphate (GMP) levels in the LC was studied using the indirect immunofluorescence technique, and the expression of soluble guanylyl cyclase with in situ hybridization. In 36 of 66 LC neurons extracellular application of SIN-1 and DEA/NO caused a hyperpolarization and a decrease in apparent input resistance. In almost 20% of neurons SIN-1 increased the firing rate. No effect could be recorded with the brain-inactive SIN-1 derivative molsidomin. The membrane permeable cGMP analogue 8-bromo-cGMP imitated the action of SIN-1. The hyperpolarizing effect of SIN-1 and DEA/NO was attenuated by preincubation with the guanylyl cyclase inhibitor ODQ. The immunohistochemical analysis revealed lack of cGMP immunostaining in non-stimulated slices, whereas SIN-1 dramatically increased this staining in about 40% of the LC neurons, and these neurons were all tyrosine hydroxylase positive, that is noradrenergic. A large proportion of the LC neurons expressed soluble guanylyl cyclase mRNA. The present and previous results suggest that NO, released from a small number of non-noradrenergic neurons in the LC, mainly has an inhibitory influence on many noradrenergic neurons, by upregulating cGMP levels via stimulation of soluble guanylyl cyclase. As nitric oxide synthase is present only in a small number of non-noradrenergic neurons (Xu et al., 1994), a few neurons may influence a large population of noradrenergic LC neurons, which in turn may control activity in many regions of the central nervous system.

Animals↗

PubMed1990

Immunohistochemical localization of 5-hydroxytryptamine, histamine and histidine decarboxylase in the rat major pelvic and coeliac-superior mesenteric ganglion.

The localization of 5-hydroxytryptamine (5-HT), histamine and histidine decarboxylase (HDC), the enzyme synthesizing histamine, was studied in the rat major pelvic and coeliac-superior mesenteric ganglia by an indirect immunofluorescence technique. Small cells (10-20 microns in diameter) exhibiting 5-HT, histamine or HDC immunoreactivities were observed in clusters or occurred as solitary cells in both ganglia. In the major pelvic ganglia, solitary histamine-immunoreactive principal neurons were also observed. Colocalization studies indicated that all 5-HT-, histamine- and HDC-immunoreactive small cells in these ganglia were labelled with tyrosine hydroxylase (TH), suggesting that they are small intensely fluorescent (SIF) cells. In the coeliac-superior mesenteric ganglia, all TH-immunoreactive SIF cells were also intensely immunoreactive for 5-HT and HDC. In the major pelvic ganglia, all TH-immunoreactive SIF cells contained 5-HT immunoreactivity, and the majority of them were also intensely immunoreactive for HDC. In both ganglia, however, only a subpopulation of TH-immunoreactive SIF cells displayed histamine immunoreactivity. The results indicate that in the rat major pelvic and coeliac-superior mesenteric ganglia, a population of catecholamine-containing SIF cells contain 5-HT and histamine suggesting a diverse role SIF cells may have in so far as modulation of ganglion transmission is concerned.

Animals↗

PubMed1986

Distribution of noradrenaline in the brain of the teleost Gasterosteus aculeatus L.: an immunohistochemical analysis.

The distribution of noradrenergic neurons in the brain of the three-spined stickleback was demonstrated with the indirect peroxidase-antiperoxidase (PAP) immunohistochemical method with antibodies against a noradrenaline-bovine serum albumin conjugate. Noradrenergic neuronal somata were exclusively located in the isthmal area of the brain stem and in the lower medulla. Noradrenergic varicose axons innervate the reticular formation, motor nuclei, and interpeduncular nucleus of the brain stem, the hypothalamus and habenular nuclei, various parts of the area dorsalis telencephali (forebrain pallium), and the olfactory bulbs. Scattered noradrenergic axons were observed in the optic tectum and in various parts of the cerebellum. It is concluded that the isthmal cell group of the stickleback is, on topological and cytoarchitectonic grounds, equivalent to the ventral portion of the locus coeruleus/subcoeruleus area of amniotes, but that its efferent connections display features characteristic both of those originating in the locus coeruleus, and in the lateral tegmental cell groups of mammals.

Animals↗

PubMed1986

Pre- and postnatal development of 5-hydroxytryptamine-immunoreactive cells in the superior cervical ganglion of the rat.

The pre- and postnatal development of 5-hydroxytryptamine (5-HT)-immunoreactive cells in the superior cervical ganglion of the rat was studied by indirect immunofluorescence method with an antibody to a conjugate of bovine serum albumin and 5-HT. The superior cervical ganglia from 11-day-old embryos to 90-day-old postnatal rats were examined. 5-HT-immunoreactive cells were first detected in the superior cervical ganglion on the 12th day of gestation. At this stage of development the ganglionic cells formed a condensed group, and most of them showed 5-HT-immunoreactivity. During later prenatal development the relative number of the 5-HT-immunoreactive cells in the ganglion decreased and most ganglionic cells appeared as non-reactive. 5-HT-immunoreactive cells showed some variation in size and fluorescence intensity during the whole prenatal development. In the ganglia of newborn rats a wide range of 5-HT-immunoreactive cell sizes was detected. Two types of 5-HT-immunoreactive cells were distinguished in the ganglia of 7- to 35-day-old rats: small (5-15 microns in diameter) cells and large (15-30 microns in diameter) cells, resembling in size small intensely fluorescent (SIF) cells and principal nerve (PN) cells, respectively. The relative number of the large 5-HT-immunoreactive cells gradually decreased after the first postnatal week, and these cells were not detected in the ganglia of 90-day-old rats. Small 5-HT-immunoreactive cells constantly formed clusters in the ganglia of postnatal rats. A marked, statistically significant increase in the density of 5-HT-immunoreactive cells occurred during the 4th postnatal week. The 5-HT-immunoreactive cells appeared in the prenatal superior cervical ganglion of the rat approximately at the same time as the cells showing formaldehyde-induced catecholamine fluorescence. Transient occurrence of the large 5-HT-immunoreactive cells during the first postnatal weeks of development indicates, that in the superior cervical ganglion of the rat, 5-hydroxytryptamine may be expressed in some developing PN cells early postnatally. After the 5th postnatal week, the large 5-HT-immunoreactive neuronal cells cannot be detected, and 5-HT is expressed only in the SIF cells.

Animals↗

PubMed1986

Glucocorticoid receptor immunoreactivity in monoaminergic neurons of rat brain.

A monoclonal antibody against the rat liver glucocorticoid receptor was used in combination with rabbit antibodies against tyrosine hydroxylase, phenylethanolamine N-methyltransferase, and 5-hydroxytryptamine to demonstrate strong glucocorticoid receptor immunoreactivity in large numbers of central monoaminergic nerve cell bodies of the male rat. The receptor immunoreactivity was predominantly located in the nucleus, whereas the tyrosine hydroxylase, phenylethanolamine N-methyltransferase, and 5-hydroxytryptamine were detected mainly in the cytoplasm. The vast majority of the noradrenergic nerve cell bodies of groups A1-A7 and of the 5-hydroxytryptaminergic cell bodies of groups B1-B9 were found to contain strong glucocorticoid receptor immunoreactivity. The majority of the phenylethanolamine N-methyltransferase-immunoreactive nerve cells of the adrenergic cell groups C1-C3 and of the dorsal subnuclei of the nucleus tractus solitarius in the medulla oblongata were also strongly immunoreactive for glucocorticoid receptor. In the midbrain dopaminergic groups A8-A10, moderately (A8, A9) to strongly (A10) glucocorticoid receptor-immunoreactive cells were found, ranging from 40 to 75% of the total population. In the hypothalamic dopaminergic cell groups, all the cells of groups A12 and A14, as well as the majority of the dopaminergic cells of the zona incerta (A13), were found to contain moderate to strong glucocorticoid receptor immunoreactivity, but none of the large dopaminergic cells of the posterior hypothalamus (A11) showed such immunoreactivity.

Amines↗

PubMed1985

Three dimensional analysis of retinal neuropeptides and amine in the chick.

Three dimensional analysis of retinal neuropeptides and monoamine-containing amacrine cells were performed on flat-mount preparations of the chick retina by using indirect immunofluorescence method. somatostatin (SOM), neurotensin (NT), leu-enkephalin (ENK), vasoactive intestinal polypeptide (VIP), substance P (SP), corticotropin releasing factor (CRF), avian pancreatic polypeptide (APP), glucagon (GLC), 5-hydroxytryptamine (5HT) and tyrosine hydroxylase (TH) were examined with specific antisera. To localize these substances in the amacrine cells, and to see in which layers their processes arborize, frozen sections were examined. There were four patterns of distribution. (1) Substances with more immunoreactive cells in the central than in the peripheral portions (SOM, NT, VIP, SP, GLC, 5HT), (2) Substances with more immunoreactive cells in the peripheral portion than in the central portion (APP), (3) Substances for which such cells were evenly distributed (TH), and (4) Substances with more immunoreactive cells in the inferior than in the superior portion (CRF). Subtypes were identified among the amacrine cells containing single peptides or monoamine.

Animals↗

PubMed1984

A method for specific transmitter identification of retrogradely labeled neurons: immunofluorescence combined with fluorescence tracing.

In the present article a method is described which allows the delineation of the projections of a single neuron as well as the identification of one or more of its chemical components. The technique is a combination of retrograde tracing and fluorescent dyes based on the work of Kuypers and collaborators and indirect immunofluorescence histochemistry as originally described by Coons and collaborators. The crucial parameters including the selection of the dyes, the injection technique and tissue processing as well as the appropriate immunohistochemical fluorescent markers and filter combinations are discussed. The method of choice involves the use of the retrogradely transported dyes Fast Blue, True Blue or Propidium Iodide, and in addition, for double labeling experiments, Diamidino Yellow or Primuline. They are combined with FITC (Propidium Iodide) or TRITC (Fast Blue, True Blue, Diamidino Yellow, Primuline) as immunofluorescence markers.

Animals↗

PubMed1983

Immunohistochemical evidence for the presence of gamma-aminobutyric acid and serotonin in one nerve cell. A study on the raphe nuclei of the rat using antibodies to glutamate decarboxylase and serotonin.

A specific and sensitive double immunocytochemical staining for the visualization of glutamate decarboxylase (GAD) and serotonin (5-HT) on the same brain section is developed. GAD is detected with specific GAD-antibodies by means of the unlabeled antibody enzyme, peroxidase anti-peroxidase, and serotonin with an antibody against the BSA-serotonin conjugate by an indirect immunofluorescent staining. The coexistence of GAD and 5-HT in the same perikaryon is demonstrated by a peroxidase reaction superimposed on fluorescent compounds. Cell bodies containing both antigens are observed in each raphe nuclei. However, the nucleus raphe dorsalis exhibits the largest number of cells containing either GAD alone or GAD and 5-HT together. An intracellular interaction between the metabolism of GABA and serotonin could be reasonably expected. The interactions between GABAergic and serotonergic systems must be thought of in terms of intracellular and/or transynaptic controls.

Animals↗

PubMed1982

The distribution and origin of serotonin-containing fibers in the septal area: a combined immunohistochemical and fluorescent retrograde tracing study in the rat.

The distribution of 5-hydroxytryptamine (serotonin, 5-HT)-containing nerve fibers and terminals in the septal area of the rat was studied by using immunohistochemistry with specific antibodies to 5-HT in combination with fluorescent retrograde tracing methods. The 5-HT innervation of the septum is heterogeneous with regard to both the morphology of individual processes and the density of distribution in different parts of the septum. Three major classes of 5-HT like immunoreactive processes can be distinguished: 1) thin, convoluted fibers with small, round or elongated varicosities; 2) thick and relatively straight fibers with few varicosities; and 3) pericellular plexuses with large varicosities in close association with perikarya in the lateral septum. Three areas of the septum receive a prominent innervation by 5-HT processes: the diagonal band of Broca, the ventral part of the lateral septum, and an area bordering the medial edge of the islands of Calleja (insula magna). Whereas the two latter areas contain dense terminal networks, the diagonal band of Broca is occupied primarily by 5-HT fibers en route to other parts of the septum. Intraseptal injections of HRP or fluorescent dyes (granular blue, propidium iodide) resulted in retrograde labeling of neuronal cell bodies in several nuclei of the brainstem which are known to contain 5-HT neurons: the dorsal raphe, the median raphe, the nucleus reticularis tegmenti pontis, the raphe pontis, and the raphe magnus. Where fluorescent retrograde tracing was performed with 5-HT immunohistochemistry on the same tissue section, a prominent 5-HT containing pathway and a non-5-HT-containing pathway from the raphe nuclei to the septum were revealed. Finally, double retrograde fluorescent labeling after injections of granular blue or propidium iodide into the septum and entorhinal area respectively of the same rat revealed extensive branching of the raphe efferents. Thus, individual raphe neurons may simultaneously connect with septum and the entorhinal area, two structures essential for normal hippocampal function.

Animals↗

PubMed1982

Identification of serotonin and non-serotonin-containing neurons of the mid-brain raphe projecting to the entorhinal area and the hippocampal formation. A combined immunohistochemical and fluorescent retrograde tracing study in the rat brain.

We have studied the localization of serotonin- and non-serotonin-containing cell bodies in the midbrain raphe nuclei that project to the entorhinal area and the hippocampal formation in the rat brain, using the technique of combined retrograde fluorescent tracing and immunohistochemistry on the same tissue section. The branching properties of these neurons were studied by retrograde double labelling using two fluorochromes which emit fluorescence with different spectral characteristics. After injections of granular blue or propidium iodide into the medial entorhinal area, retrogradely-labelled cells were found situated bilaterally in the caudal half of the dorsal raphe nucleus, the medial part of the median raphe and throughout the rostrocaudal extension of the nucleus reticularis tegmentipontis. Injections placed successively more laterally in the entorhinal area labelled progressively less cells contralaterally in the dorsal raphe and the reticular tegmental nucleus of the pons. After fluorochrome injections into the dorsal part of the hippocampal formation, retrogradely-labelled cells were found in the caudal part of the dorsal raphe, in the peripheral part of the median raphe and to a minor extent in the medial part of this nucleus, but not in the nucleus reticularis tegmentipontis. The experiments with double retrograde fluorescent tracing showed that the raphe nuclei do not send bilateral projections to the entorhinal area in spite of the fact that many of these cells are located contralateral to the injected hemisphere in single labelling experiments. Injections of the fluorochromes into the entorhinal area and hippocampal formation showed that at least 10% of the raphe cells project to both areas simultaneously. Analysis of sections incubated with antiserum to serotonin showed that a majority of the retrogradely-labelled versus serotonin-immunoreactive cells was found to vary within different parts of the individual raphe nuclei: the ventromedial part of the dorsal, the medial part of the median and the nucleus reticularis tegmentipontis being the highest. The findings indicate that both serotonin- and non-serotonin-containing neurons in the raphe innervate the hippocampal region, that these projections may be crossed but not bilateral, and that the same neuron in the raphe may influence the neural activity in the entorhinal area and the hippocampus simultaneously.

Animals↗