Intracerebral microinjection of interleukin-4/interleukin-13 reduces ß-amyloid accumulation in the ipsilateral side and improves cognitive deficits in young amyloid precursor protein 23 mice.

We previously reported that the anti-inflammatory cytokine interleukin (IL)-4 induced selective clearance of oligomeric ß-amyloid (Aß(1-42)) in rat primary type 2 microglial cells. For the present study, we investigated whether IL-4 and IL-13 could activate microglial cells to induce Aß clearance in vivo and improve cognitive deficits in APP23 mice, which are amyloid precursor protein transgenic mice. We administered an intracerebral microinjection of a mixture of IL-4 and IL-13 or of saline vehicle into one hemisphere of APP23 mice and their wild-type littermates, 4.5 and 9 months old, after which we evaluated the effects of these treatments on spatial learning and memory by Morris Water Maze test and on accumulated amounts of Aß. The cytokine injection significantly improved memory deficits of 4.5-month-old APP23 mice, but did not do so in 9-month-old APP23 mice, even though similar Aß reductions were observed in both age groups of APP23 mice in the ipsilateral neocortex. The cytokine injection improved memory impairment of 9-month-old wild-type (WT) mice in the probe trial. Immunohistochemical analysis of the 4.5-month-old APP23 mice revealed the presence of increased numbers of microglial cells at 2 days after the cytokine injection. In addition to induced CD36 expression in the activated microglia, increased expression of neprilysin, mainly in neurons, suggested that the cytokines improved the cognitive deficits via degradation and clearance of intra- and extraneuronal Aß peptides, of buffer-extractable nonplaque form. Double immunostaining also revealed that most of the activated microglia had the M2-like phenotype. This unique mechanism of IL-4/IL-13-induced clearance of Aß may provide an additional strategy to prevent and/or cure Alzheimer's disease at early stage.

Noradrenergic excitation of a subpopulation of GABAergic cells in the basolateral amygdala via both activation of nonselective cationic conductance and suppression of resting K+ conductance: a study using glutamate decarboxylase 67-green fluorescent protein knock-in mice.

GABAergic interneurons play central roles in the regulation of neuronal activity in the basolateral nucleus of the amygdala (BLA). They are also suggested to be the principal targets of the brainstem noradrenergic afferents which are involved in the enhancement of the BLA-related memory. In addition, behavioral stress has been shown to impair noradrenergic facilitation of GABAergic transmission. However, the noradrenaline (NA) effects in the BLA have not been differentiated among medium- to large-sized GABAergic neurons and principal cells, and remain to be elucidated in terms of their underlying mechanisms. Glutamate decarboxylase 67 (GAD67) is a biosynthetic enzyme of GABA and is specifically expressed in GABAergic neurons. To facilitate the study of the NA effects on GABAergic neurons in live preparations, we generated GAD67-green fluorescent protein (GFP) knock-in mice, in which GFP was expressed under the control of an endogenous GAD67 gene promoter. Here, we show that GFP was specifically expressed in GABAergic neurons in the BLA of this GAD67-GFP knock-in mouse. Under whole-cell patch-clamp recordings in vitro, we identified a certain subpopulation of GABAergic neurons in the BLA chiefly on the basis of the electrophysiological properties. When depolarized by a current injection, these neurons, which are referred to as type A, generated action potentials at relatively low frequency. We found that NA directly excited type-A cells via alpha1-adrenoceptors, whereas its effects on the other types of neurons were negligible. Two ionic mechanisms were involved in this excitability: the activation of nonselective cationic conductance and the suppression of the resting K+ conductance. NA also increased the frequency of spontaneous IPSCs in the principal cells of the BLA. It is suggested that the NA-dependent excitation of type-A cells attenuates the BLA output for a certain period.

Differential distribution of vesicular glutamate transporters in the rat cerebellar cortex.

The chemical organization of excitatory axon terminals in the rat cerebellar cortex was examined by immunocytochemistry and in situ hybridization histochemistry of vesicular glutamate transporters 1 and 2 (VGluT1 and VGluT2). Chemical depletion of the inferior olivary complex neurons by 3-acetylpyridine treatment almost completely removed VGluT2 immunoreactivity from the molecular layer, leaving VGluT1 immunoreactivity apparently intact. On the other hand, neuronal deprivation of the cerebellar cortex by kainic acid injection induced a large loss of VGluT1 immunoreactivity in the molecular layer. In the cerebellar granular layer, both VGluT1 and VGluT2 immunoreactivities were found in mossy fiber terminals, and the two immunoreactivities were mostly colocalized in single-axon terminals. Signals for mRNA encoding VGluT2 were found in the inferior olivary complex, and those for VGluT1 and VGluT2 mRNAs were observed in most brainstem precerebellar nuclei sending mossy fibers, such as the pontine, pontine tegmental reticular, lateral reticular and external cuneate nuclei. These results indicate that climbing and parallel fibers selectively use VGluT2 and VGluT1, respectively, whereas mossy fibers apply both VGluT1 and VGluT2 together to accumulate glutamate into synaptic vesicles. Since climbing-fiber and parallel-fiber terminals are known to make depressing and facilitating synapses, respectively, VGluT1 and VGluT2 might have distinct properties associated with those synaptic characteristics. Thus, it would be the next interesting issue to determine whether mossy-fiber terminals co-expressing VGluT1 and VGluT2 show synaptic facilitation or depression.

In vivo transduction of central neurons using recombinant Sindbis virus: Golgi-like labeling of dendrites and axons with membrane-targeted fluorescent proteins.

A new recombinant virus which labeled the infected neurons in a Golgi stain-like fashion was developed. The virus was based on a replication-defective Sindbis virus and was designed to express green fluorescent protein with a palmitoylation signal (palGFP). When the virus was injected into the ventrobasal thalamic nuclei, many neurons were visualized with the fluorescence of palGFP in the injection site. The labeling was enhanced by immunocytochemical staining with an antibody to green fluorescent protein to show the entire configuration of the dendrites. Thalamocortical axons of the infected neurons were also intensely immunostained in the somatosensory cortex. In contrast to palGFP, when DsRed with the same palmitoylation signal (palDsRed) was introduced into neurons with the Sindbis virus, palDsRed neither visualized the infected neurons in a Golgi stain-like manner nor stained projecting axons in the cerebral cortex. The palDsRed appeared to be aggregated or accumulated in some organelles in the infected neurons. Anterograde labeling with palGFP Sindbis virus was very intense, not only in thalamocortical neurons but also in callosal, striatonigral, and nigrostriatal neurons. Occasionally there were retrogradely labeled neurons that showed Golgi stain-like images. These results indicate that palGFP Sindbis virus can be used as an excellent anterograde tracer in the central nervous system.

Guidance of glial precursor cell migration by secreted cues in the developing optic nerve.

Oligodendrocyte precursors are produced in restricted foci of the germinative neuroepithelium in embryo brains and migrate to their sites of function, while astrocytes are produced in a wider area in the neuroepithelium. We investigated the guidance mechanisms of glial precursor (GP) cell migration in the optic nerve. GP cell migration in newborn rat optic nerve was monitored by the UV-thymine-dimer (TD) method. A double labeling study using NG2 and TD revealed that many of these in vivo migrating cells were NG2 positive, while some of them with large TD-positive nuclei were NG2 negative. An in vitro cell migration study using optic nerve with chiasma and/or eyeball tissue revealed that the GP cells migrated under the guidance of repulsive cues secreted from the optic chiasma. We detected the expression of netrin 1 and Sema3a in the optic chiasma, and that of Unc5h1 and neuropilin 1 in the optic nerve. Co-culture experiments of the optic nerve with cell clusters expressing guidance cues revealed that the migrating GP cells in the optic nerve were heterogeneous. Netrin 1 repelled a subtype of NG2-positive and PLP-positive GP cells with small nuclei. Sema3a repelled a subtype of GP cells with large nuclei.

Radial glia is a progenitor of neocortical neurons in the developing cerebral cortex.

Neocortical neurons are produced by cell division of neural stem cells in the ventricular zone of the cerebral cortex. We investigated the production of neurons by infecting neuroepithelial cells with a modified GFP-recombinant adenovirus. The adenovirus DNA is inherited by only one daughter cell at each cell division and travels one way from the progenitor to the progeny. Since the ventricular zone (VZ) of the embryo neocortex expressed an adenovirus receptor, CAR ubiquitously, morphology and cell-lineage of cells in the VZ could be revealed by the adenovirus infection. Radial glias, cells with a bipolar shape, and spherical cells were found as modified-GFP-positive (mGFP+) in the VZ. The bipolar cells (radial cells) had a radial process not in contact with the pia mater and a growth-cone-like structure at the edge of their radial process, while the radial glias had a process spanning all the cortical layers. Ten hours after viral infection, most mGFP+ cells were radial cells. In the following 8 h, the percentage of mGFP+ radial glias in mGFP+ neocortical cells increased from 18 to 50%, while that in radial/spherical cells decreased from 75 to 19%. The radial glias often divided asymmetrically and produced spherical cells and neuronal precursors. The spherical cells seemed to become radial cells by extending a radial process. The spherical cells, radial cells and radial glias seemed to constitute a proliferating cell cycle during which postmitotic neuronal precursors are produced. The neuronal precursors that inherited the radial processes migrated radially and developed into neocortical neurons. Four days after the viral infection, 97% of mGFP+ cells were neocortical neurons. Here, we propose that the radial glia is a progenitor of neocortical neurons, and that a significant number of radially migrating neurons is guided by their own radial processes connected to the pia mater.

Distribution of preganglionic terminals in the cervical sympathetic ganglia detected by the expression of c-Fos like protein after electric stimulation of the ventral root.

To determine the segmental relationship between the upper thoracic spinal cord and cervical sympathetic ganglia, we observed the distribution pattern of postganglionic cells which expressed c-Fos like protein, one of the products of immediate early genes, after electrical stimulation of ventral roots at the T1-T3 spinal segments. We recognized a clear segmental arrangement of postganglionic cells in the stellate ganglion along its rostrocaudal direction corresponding to the segmental arrangement of preganglionic neurons in the spinal cord. That is, postganglionic neurons which expressed c-Fos like protein after stimulation of the T1 ventral root were distributed in the middle region of the stellate ganglion in the rostrocaudal direction. The c-Fos like protein-positive neurons after stimulation of the T2 ventral root were distributed in a more caudal region of the stellate ganglion than after T1 ventral root stimulation. C-Fos like protein-positive neurons after stimulation of the T3 ventral root were mainly situated in a more caudal region of the stellate ganglion than after T2 ventral root stimulation. There was, however, no segmental relationship between the upper thoracic levels of the spinal cord and superior cervical ganglion in the rostrocaudal direction. These results indicate that the segmental innervation of the upper thoracic spinal cord exists in the stellate ganglion, but not in the superior cervical ganglion.

Cell migration from the corticostriatal angle to the basal telencephalon in rat embryos.

Lateral cortical stream (LCS) has been described as a flow of cell migration found in the lateral cortex of the embryonic telencephalon of mammals. The destinations of the cell migration were reported as the ventrolateral neocortex, the pyriform cortex, endopyriform nucleus and the claustrum. At the same time, however, other destinations of LCS have been suggested in the ventral telencephalon. Therefore, we investigated the additional destinations of the LCS using a combination of several molecular biological techniques. Using an expression vector of modified green fluorescent protein (GFP) introduced into the ventricular zone (VZ) around the corticostriatal angle, both tangential and radial cell migration was revealed in the LCS. The radial cell migration in the LCS supplied cells to the ventro-lateral neocortex, pyriform cortex and to the level of the lateral olfactory tract. In the second experiment, we injected COS-1 cells transfected with a Sema3A expression vector into one side of the neocortex. The cell supply to the destination of the LCS ceased due to the formation of a large necrosis in the LCS, which was triggered by the Sema3A-COS cell injection, and the dense cell layer in the olfactory tubercle shrunk on the side where COS cells were injected. These data indicated that the majority of neurons in the dense cell layer of the olfactory tubercle reached this point through the LCS. One of the origins of the cells in the LCS would be in the corticostriatal angle.

Neurons in Golgi-stain-like images revealed by GFP-adenovirus infection in vivo.

Neurons in the adult brain have a very complex morphology with many processes, including tremendously long axons. Since dendrites and axons play key roles in the input and output of neural information, respectively, the visualization of complete images of these processes is necessary to reveal the mechanism of neural information processing. Here we made a recombinant adenovirus vector which encodes green fluorescent protein (GFP) tagged with a palmitoylation site, a membrane-targeting signal, produced specific antibodies to GFP, and used them as probes for staining the nervous system. In the neocortex, after injection of the recombinant virus and immunoperoxidase staining with the antibodies, many different types of cells were labeled in a Golgi stain-like fashion. Although the number of labeled cells varied depending on the amount of virus injected, the recombinant virus was considered to be infectious to cortical neurons of all cell types without selectivity. In contrast, the viral infection in the cerebellar cortex and superior cervical ganglion showed some selectivity toward the cell type. It is expected that this recombinant virus will be a useful tool for the morphological analysis of neuronal connections, especially the analysis of microcircuitry in the cerebral cortex.

Cell migration from the ganglionic eminence to the neocortex investigated by labeling nuclei with UV irradiation via a fiber-optic cable.

Recent studies have shown that the ganglionic eminence is one of the sources of tangentially migrating cells in the developing neocortex. Since the migration of the DiI-labeled cells from the ganglionic eminence to the neocortex was not monitored by videomicroscopy in these reports, we devised a novel method to study cell migration in vitro and in vivo. The new method involves ultraviolet (UV) irradiation of the cells through a fiber-optic cable and subsequent identification of the irradiated cells on the basis of the formation of thymine dimers in the nuclei. First, we tested the new method (UV-thymine dimer-labeling method) by applying it to monitor the cell migration of neuronal precursor cells in the rostral migratory stream in the neonatal rat telencephalon. In vitro, UV irradiation for 1 s through the fiber-optic cable resulted in the formation of sufficient thymine dimers as to allow immunohistochemical detection after 6 h of incubation; a significant proportion of the irradiated cells continued to migrate in the same direction and at the same speed as those before irradiation. There was no significant difference in the cell migration distance over 6 h between cells exposed and not exposed to the UV irradiation in vitro. In vivo, this method revealed that three times as many cells in the subventricular zone of the olfactory bulb migrated rostrally as caudally. The new method also allowed us to measure the speed of cell migration, which was estimated to be about 70 microm/h at the maximum in the rostral direction. After these examinations of reliability of the method, we applied it to the rat embryo brain. One day after UV irradiation of the ganglionic eminence, labeled migrating cells were found in the striatum, in the internal capsule, and in the intermediate zone of the neocortex. The observation period of cell migration to the neocortex was extended by the use of a xeroderma pigmentosum group A gene mutant mouse, which lacked an ability to remove thymine dimer from the UV-irradiated nuclei. Two days after the UV irradiation, labeled migrating cells from the ganglionic eminence of the mutant mouse embryos were found both in the cortical plate and in the intermediate zone of the neocortex.

Development of afferent fiber lamination in the infrapyramidal blade of the rat dentate gyrus.

In the rat dentate gyrus, the lateral perforant path, the medial perforant path, and the major part of the hilar projection to the molecular layer share the lamination domain, mainly in the outer one-third of the molecular layer, the middle one-third, and the inner one-third, respectively. To reveal the order of the afferent fiber lamination and to have an indication of how the synaptic sites on dendrites are determined, we investigated the ontogeny of afferent fiber lamination in the dorsal hippocampus by injecting 1, 1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) into the entorhinal cortex and hippocampus in vivo. Fibers from the contralateral hilar region were found under the pia mater of the infrapyramidal blade at postnatal day 3 (P3), whereas the entorhinal afferent fibers were absent in the infrapyramidal blade. Then the medial and the lateral perforant path appeared under the pia mater in the infrapyramidal blade as riding on top of the preexisting laminae by P7 and by P11, respectively. Based on the established knowledge that most entorhinal layer II neurons simultaneously innervate both the suprapyramidal blade and infrapyramidal blade by branching, it is assumed that the medial and lateral perforant path in the suprapyramidal blade await an appropriate timing for sprouting of interstitial branches into the infrapyramidal blade. The granule cells in the infrapyramidal blade had dendritic growth cones by P11. Calretinin immunohistochemistry revealed Cajal-Retzius cells in the infrapyramidal blade even at P14. Under the pia mater, axon growth cones of ingrowing afferent fibers may interact with the dendritic growth cones or the Cajal-Retzius cells, and determines the synaptic sites on the granule cell dendrites.

Expression of Hex mRNA in early murine postimplantation embryo development.

The onset of Hex expression and its role in early murine development was analyzed using in situ hybridization. Hex mRNA was first detected in the chorion of the ectoplacental cavity and weakly at the visceral endoderm of the future yolk sac at embryonic age (E) 7.5. Expression in embryonic tissues was detected exclusively in the hepatic anlage and thyroid primordium at E 9.5. At E 12.5 and E 15.5, Hex expression persisted in the fetal liver and thyroid, and was also detected in the fetal lung. These results suggest that Hex has its role in differentiation and/or organogenesis of several embryonic tissues.

Organization of the entorhinal projection to the rat dentate gyrus revealed by Dil anterograde labeling.

It has been suggested that the entorhino-hippocampal circuit is involved in memory formation. To investigate the way that associative memory is elaborated in the circuit, the entorhino-dentate projection was studied with the fluorescent lipophilic tracer Dil. We investigated the projection originating in the dorsal part of the entorhinal cortex by injecting Dil along the rhinal sulcus. Anterograde fluorescent labeling allowed us to examine sections of the sample with a confocal microscope or in wholemount preparations with a fluorescence microscope. Quantitative analysis of the distribution of the Dil-labeled perforant path by confocal microscopy was performed in the septal one third level of the hippocampus. The analysis confirmed that the topographical map along the mediolateral dimension of the entorhinal cortex was transferred to the proximodistal level (from the inner one third to the edge of the molecular layer) of the granule cell dendrites in a gradually shifting manner. The fiber profile observed after lateral entorhinal injection was thick in the suprapyramidal blade and thin in the infrapyramidal blade. The fiber profile observed after medial entorhinal injection was thin in the suprapyramidal blade and thick in the infrapyramidal blade. Fluorescence microscopic observation of wholemount preparations showed that projections from the Dil injection site were distributed wider than half the dentate gyrus in the longitudinal direction. In transverse sections, the range of the labeled fiber distribution was confirmed to be more than two thirds of the dentate gyrus in the same direction regardless of the mediolateral level of the injection site. It has been suggested that the dorsoventral axis of the entorhinal cortex is represented in the septotemporal levels of the dentate gyrus, but that the topographical correspondence might be weak and vague. Although our investigation was limited to the projection from the dorsal entorhinal cortex to the dorsal part of the dentate gyrus, we conclude that the widely distributed projection covers the dentate gyrus in a nontopographic manner.

Origin and route of tangentially migrating neurons in the developing neocortical intermediate zone.

Neuroblasts produced in the ventricular zone of the neocortex migrate radially and form the cortical plate, settling in an inside-out order. It is also well known that the tangential cell migration is not negligible in the embryonic neocortex. To have a better understanding of the tangential cell migration in the cortex, we disturbed the migration by making a cut in the neocortex, and we labeled the migrating cells with 1,1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) in vivo and in vitro. We also determined the birth dates of the cells. Disturbance of tangential cell migration caused an accumulation and disappearance of microtubule-associated protein 2 immunoreactive (MAP2-IR) cells on the ventral and dorsal side of the cut, respectively, which indicated that most of the MAP2-IR cells in the intermediate zone (IZ) were migrating toward the dorsal cortex. The DiI injection study in vivo confirmed the tendency of the direction of cell migration and suggested the origin of the cells to be in the lateral ganglionic eminence (LGE). DiI injection into the LGE in vitro confirmed that the LGE cells cross the corticostriatal boundary and enter the IZ of the neocortex. The migrating cells acquired multipolar shape in the IZ of the dorsal cortex and seemed to reside there. A 5-bromo-deoxyuridine incorporation study revealed that the migrating MAP2-IR cells in the IZ were early-generated neurons. We concluded that the majority of tangentially migrating cells were generated in the LGE and identified as a distinct population that was assumed not to have joined the cortical plate.

Arborization pattern of sympathetic preganglionic axons in the rat superior cervical and stellate ganglia.

Anterograde labeling technique with Phaseolus Vulgaris leucoagglutinin (PHA-L) was employed to observe how a single preganglionic axon arborizes in the superior cervical ganglion (SCG) and stellate ganglion (STG) of rats. PHA-L was injected into the intermediolateral nucleus of the spinal cord at the middle point between segments T1 and T2, and labeled axons were detected immunohistochemically in serial sections. We traced and drew three preganglionic axons over their full length in the SCG and STG. In SCG, the labeled axons bifurcated repeatedly and extended to a length of 600-700 microns in the rostrocaudal direction, and about 200 microns in the transverse direction. These three preganglionic axons made 11, 14 and 11 dense terminal plexus regions along their trajectory. The pattern of the most dense terminal plexus corresponded to the pericellular type dendritic plexus, one of the plexus patterns of dendritic collaterals of SCG neurons. In the STG, the extent of axonal arborization was more variable than that in the SCG, ranging from 400 to 800 microns in the rostrocaudal direction and about 400 microns in the transverse direction. The three analyzed axons made 21, 19 and 20 dense terminal plexus regions along their trajectory, with a similar pattern to those in SCG. These results indicated that there might be a columnar or ellipsoidal organization of postganglionic neurons which are innervated by single preganglionic axons.

A procedure for in situ hybridization combined with retrograde labeling of neurons: application to the study of cell adhesion molecule expression in Dil-labeled rat pyramidal neurons.

We have devised a simple method that combines retrograde labeling of projecting neurons and in situ hybridization histochemistry to examine mRNA expression in the retrogradely labeled neurons. First, projecting neurons were retrogradely labeled in vivo by injection of the lipophilic neuronal tracer Dil. The fluorescence of the labeled neurons in the brain slices was photoconverted into stable DAB precipitate by green light illumination. The slices were cut into thinner sections and processed for detection of specific mRNA by in situ hybridization. Using this highly sensitive method, we demonstrate here that the corticospinal tract neurons in newborn rats express mRNA for the cell adhesion molecule L1. TAG-1 mRNA was not detected in these neurons. Therefore, the present method provides an important tool to study the molecular expression of projection neurons during the development of neuronal circuitry.

Intermediate zone and intermediate zone neurons. Do IZ neurons acts as pioneers for layer V neurons in rats?

It is assumed that pioneer neurons will predominantly show path finding ability, and that other neurons only follow the pioneer fibers. Here, tracers were injected into the newborn rat spinal cord followed by a search for labeled cells other than those in layer V. In all cases, neurons in the lower intermediate zone (IZ) of the dorsomedial cortex were retrogradely labeled. The IZ neurons have several characteristics required for pioneer neurons and project to the spinal cord first. Thus, the IZ neurons may function as pioneer neurons of the corticospinal projection and help in the formation of the permanent tract in rats.

Distribution of vagal preganglionic neurons in the rat brain innervating thoracic and abdominal organs revealed by retrograde DiI tracing.

To investigate the distribution and number of preganglionic neurons which regulate motility and secretion in thoracic and abdominal organs in the vagal parasympathetic nervous system, the neuronal tracer DiI was injected into the organs and the distribution of retrogradely labeled neurons was examined in the rat brainstem. The stomach received the vast majority of efferent projections from the dorsal motor nucleus of the vagus nerve (DMV). The cecum and the duodenum also received projections from the DMV, but they originated from a smaller number of preganglionic neurons. Preganglionic neurons projecting to the stomach occupied the middle part of the DMV, those projecting to the cecum occupied the lateral part of the DMV, and those projecting to the duodenum were found in the medial edge of the DMV. The ventral and dorsal sides of the stomach wall were innervated by the left and right vagus nerves, respectively. However, immediately after passing the boundary between the stomach and duodenum, the left and right vagal nerve fibers mixed in the ventral and dorsal walls of the distal gastrointestinal tract. The nucleus ambiguous is a mixture of parasympathetic preganglionic neurons and motoneurons. In this study, we revealed that the major targets of these preganglionic neurons were the lungs and other thoracic organs.

Effects of brain stem parabrachial activation on receptive field properties of cells in the cat's lateral geniculate nucleus.

1. The lateral geniculate nucleus is the primary thalamic relay for the transfer of retinal signals to the visual cortex. Geniculate cells are heavily innervated from nonretinal sources, and these modify retinogeniculate transmission. A major ascending projection to the lateral geniculate nucleus arises from cholinergic cells in the parabrachial region of the brain stem. This is an important pathway in the ascending control of arousal. In an in vivo preparation, we used extracellular recordings to study the effects of electrical activation of the parabrachial region on the spontaneous activity and visual responses of X and Y cells in the lateral geniculate nucleus of the cat. 2. We studied the effects of two patterns of parabrachial activation on the spontaneous activity of geniculate cells. Burst stimulation consisted of a short pulse at high frequency (16 ms at 250 Hz). Train stimulation was of longer duration at lower frequency (e.g., 1 s at 50 Hz). The firing rate of almost all geniculate cells was enhanced by either pattern of stimulation. However, the burst pattern of stimulation elicited a short, modulated response with excitatory and inhibitory epochs. We found that the different epochs could differentially modulate the visual responses to drifting gratings. Thus the temporal alignment of the brain stem and visual stimuli was critical with burst stimulation, and varied alignments could dramatically confound the results. In comparison, the train pattern of stimulation consistently produced a relatively flat plateau of increased firing, after a short initial period of more variable effects. We used the less confounding pattern of train stimuli to study the effects of parabrachial activation on visual responses. 3. Our main emphasis was to examine the parabrachial effects on the visual responses of geniculate cells. For most visual stimuli, we used drifting sine wave gratings that varied in spatial frequency; these evoked modulated responses from the geniculate cells. Parabrachial activation enhanced the visual responses of almost all geniculate cells, and this enhancement included both increased depth of modulation and greater response rates. 4. Our results were incorporated quantitatively into a difference-of-Gaussians model of visual receptive fields in order to study the parabrachial effects on the spatial structure of the receptive field. This model fit our data well and provided measures of the response amplitude and radius of the receptive field center (Kc and Rc, respectively) and the response amplitude and radius of the receptive field surround (Ks and Rs, respectively).(ABSTRACT TRUNCATED AT 400 WORDS)

Morphology of physiologically identified retinal X and Y axons in the cat's thalamus and midbrain as revealed by intraaxonal injection of biocytin.

Prior morphological studies of individual retinal X and Y axon arbors based on intraaxonal labeling with horseradish peroxidase have been limited by restricted diffusion or transport of the label. We used biocytin instead as the intraaxonal label, and this completely delineated each of our six X and 14 Y axons, including both thalamic and midbrain arbors. Arbors in the lateral geniculate nucleus appeared generally as has been well documented previously. Interestingly, all of the labeled axons projected a branch beyond thalamus to the midbrain. Each X axon formed a terminal arbor in the pretectum, but none continued to the superior colliculus. In contrast, 11 of 14 Y axons innervated both the pretectum and the superior colliculus, one innervated only the pretectum, and two innervated only the superior colliculus. Two of the Y axons were quite unusual in that their receptive fields were located well into the hemifield ipsilateral with respect to the hemisphere into which they were injected. These axons exhibited remarkable arbors in the lateral geniculate nucleus, diffusely innervating the C-laminae and medial interlaminar nucleus, but, unlike all other X and Y arbors, they did not innervate the A-laminae at all. In addition to these qualitative observations, we analyzed a number of quantitative features of these axons in terms of numbers and distributions of terminal boutons. We found that Y arbors contained more boutons than did X arbors in both thalamus and midbrain. Also, for axons with receptive fields in the contralateral hemifield (all X and all but two Y axons), 90-95% of their boutons terminated in the lateral geniculate nucleus; the other two Y axons had more of their arbors located in midbrain.