Lucifer Yellow Injection Research Articles

Localization of the myomodulin-like immunoreactivity in the leech CNS.

The distribution of myomodulinlike immunoreactivity in the leech CNS was determined using an antiserum raised against Aplysia myomodulin. Segmental ganglia contained approximately 60 immunoreactive neurons. In addition, numerous fibers containing immunoreactive varicosities were found throughout the neuropil. Using a combination of Lucifer Yellow injections and immunocytochemistry, we identified neurons including the anterior Pagodas (AP), annulus erector (AE), motor neurons, Leydig, longitudinal muscle motoneurons (L), S cells, and coupling interneurons, all of which are active during the touch-elicited shortening reflex. FMRF-amide-like immunoreactivity in three of these cells (L, AP, and AE) was previously demonstrated. Specific staining for myomodulin was abolished by preadsorption of the antiserum with synthetic myomodulin, but not with FMRF-amide. These results suggest a potential role for myomodulin in both intrinsic and extrinsic modulation of the leech touch-elicited shortening reflex. Further, it is possible that several neurons mediating this reflex contain multiple neuromodulatory peptides.

Localization of the myomodulin‐like immunoreactivity in the leech CNS

The distribution of myomodulinlike immunoreactivity in the leech CNS was determined using an antiserum raised against Aplysia myomodulin. Segmental ganglia contained approximately 60 immunoreactive neurons. In addition, numerous fibers containing immunoreactive varicosities were found throughout the neuropil. Using a combination of Lucifer Yellow injections and immunocytochemistry, we identified neurons including the anterior Pagodas (AP), annulus erector (AE), motor neurons, Leydig, longitudinal muscle motoneurons (L), S cells, and coupling interneurons, all of which are active during the touch-elicited shortening reflex. FMRF-amide-like immunoreactivity in three of these cells (L, AP, and AE) was previously demonstrated. Specific staining for myomodulin was abolished by preadsorption of the antiserum with synthetic myomodulin, but not with FMRF-amide. These results suggest a potential role for myomodulin in both intrinsic and extrinsic modulation of the leech touch-elicited shortening reflex. Further, it is possible that several neurons mediating this reflex contain multiple neuromodulatory peptides. © 1996 John Wiley & Sons, Inc.

Monosynaptic input from cutaneous sensory afferents to fin motoneurons in lamprey.

The sensory control of lamprey dorsal fin motoneurons was studied by using paired intracellular recordings combined with a morphological analysis. Dorsal cells innervating the skin of the dorsal fin and fin motoneurons were retrogradely labeled by injecting fluoresceincoupled dextran amines into the dorsal fin. Labeled motoneurons and dorsal cells showed close appositions, suggesting that the dorsal cells innervating the fin region make monosynaptic connections with fin motoneurons. By using conventional electrophysiological criteria, monosynaptic excitatory connections were found between fin dorsal cells and fin motoneurons. In addition, Lucifer yellow injection followed by confocal three-dimensional (3-D) reconstructions of monosynaptically connected pairs, revealed close apposition between dorsal cell axons and the distal dendrites of fin motoneurons. Each fin motoneuron received monosynaptic excitatory input from at least four different afferents. The amplitude of the monosynaptic excitatory postsynaptic potential (EPSP)s was reduced by administration of the N-methyl-D-aspartate (NMDA) receptor antagonist DL,2 amino-5-phosphovaleric acid (APV). Sensory stimulation could also elicit di- or oligosynaptic inhibitory postsynaptic potential (IPSP)s, which were blocked by the glycine antagonist strychnine, resulting in the appearance of large monosynaptic EPSPs, which could induce action potentials.

Blue-Cone Horizontal Cells in the Retinae of Horses and OtherEquidae

The morphology of horizontal cells chiefly of the horse, but also of asses, mules, and a zebra, has been examined by Lucifer yellow injections into lightly fixed retinae and by immunocytochemistry. In common with other mammals, equids have a B-type horizontal cell, i.e., a cell with dendrites synapsing with cones and possessing a single axon synapsing with rods. Most mammalian retinae have a further type of horizontal cell, the A-type, also synapsing with cones but without an axon. The second type of horizontal cell in equids also has no axon; otherwise, it is most unusual. Compared with other mammalian A-type cells, it has a vary large dendritic field, both absolutely and relative to the dendritic fields of B-type cells. The dendrites are fine and sparsely branching. Their most striking feature is that they bear a low density of irregularly spaced synaptic terminal aggregates, suggesting their cone contacts are selective. Immunolabelling of S (blue)-cones in horse retina showed that they comprise, depending on retinal location, 10-25% of the cone population. For a single horse A-type cell, it is shown that 44 of its 45 terminal aggregates are congruent with the pedicles of S-cones. Immunostaining with a calbindin antibody demonstrated that each type of horizontal cell forms an independent regular mosaic. The density ratio of B- to A-type cells varied between 5 and 10. This is the first demonstration in a mammalian retina of a horizontal cell type with a direct input exclusively from S-cones.

Synaptic potentials recorded from sea anemone muscle cells in situ.

Three pulse types, A,B, and C, can be recorded from column ring preparations of the sea anemone Calliactis parasitica (Couch). Lucifer Yellow injection confirmed that the recording sites are intracellular in endodermal myoepithelial cells. Type C pulses are spontaneously active depolarizing pulses, arising from a resting potential of around -60mV, and are usually < 5 mV in amplitude. When the cells were voltage-clamped these pulses reversed at around 0 mV membrane potential; therefore, we conclude that they are synaptic potentials. In some cells an increase in frequency of type C pulses accompanied contraction of the circular muscle field. Inward currents in these myoepithelial cells are carried by Ca2+, and although Na(+)-free solutions did not affect inward currents they did eliminate type C pulses, i.e., these synaptic potentials may result from activity of Na(+)-dependent nerve cells.

Synaptically released histamine increases dye coupling among vasopressinergic neurons of the supraoptic nucleus: mediation by H1 receptors and cyclic nucleotides

Activating direct olfactory (glutamatergic) inputs to supraoptic nucleus (SON) neurons increases interneuronal coupling in slices from lactating but from not virgin or male rats. Studied here were influences on coupling of another monosynaptic input to SON, the histaminergic tuberomammillary nucleus (TM) projection, activation of which selectively excites phasically firing (putative vasopressin) cells. Effects of TM stimulation and its possible downstream consequences on Lucifer yellow (LY) dye coupling among putative vasopressin cells were determined in male rat SONs. In unstimulated slices, 12 LY injections (1 cell/SON) yielded eight single and four pairs of coupled neurons. In slices in which TM was stimulated for 10 min at 10 Hz, 13 injections yielded 4 single and 28 coupled cells, with groups of 2 to 4 cells coupled to the injected neuron, a threefold increase in the number of coupled cells per injection (p < 0.02). Bathing slices in medium containing 10 microM pyrilamine (H1 antagonist) blocked this stimulation-induced coupling increase, suggesting mediation by activation of guanylate cyclase-cGMP to which H1 receptors often are linked . Bathing slices in medium containing 0.5-1 mM 8-bromo-cGMP yielded results similar to those of TM stimulation, a 2.5-fold increase over control in the number of coupled cells per injection. Effects of TM stimulation on coupling also were blocked by bathing slices in a guanylate cyclase inhibitor (10 microM LY83583). In contrast to cGMP, 1 mM 8-bromo-cAMP significantly reduced coupling. We conclude that synaptically released histamine increases coupling via cGMP-dependent mechanisms.

Confocal Laser Scanning Microscopy and 3-D Reconstructions of Neuronal Structures in Human Brain Cortex

Human brain material was studied with Lucifer yellow (LY) microinjections, indirect Texas red immunofluorescence, and confocal laser scanning microscopy (CLSM). The scanned images were transferred to a Silicon Graphics (SG) IRIS computer equipped with software for reconstructing the 3-D architecture of cells. By employing dual channel CLSM (Bio-Rad MRC 600), LY-injected cells and Texas red immunofluorescence could be studied simultaneously. Autopsy material with 2 to 48-h postmortem delays (6 control and 2 Rett's syndrome cases) as well as biopsy material (14 cases with therapy-resistant partial epilepsy-TRPE-undergoing neurosurgery) were used. In each specimen, 100-200 pyramidal and nonpyramidal neurons were visualized by LY microinjection. Single neurons were imaged and 2-D reconstructions of each neuron were made using z-projections of serial optical images; 3-D reconstructions and rotations were computed using the SG workstation, with VoxelView software from Vital Images (UK), and stored in a "neuronal library" on laser or magnetic optical disks. In Rett's syndrome cases and in patients with TRPE various abnormalities in the dendritic geometry of pyramidal and nonpyramidal cells have been found. The combination of LY injections with immunofluorescence allows the investigation of transmitter-related substances around the LY-injected cells. Using antibodies to synaptic vesicle proteins, presynaptic elements docking onto individual spines have been demonstrated. This approach may contribute to the understanding of different neurological and psychiatric disorders and may be useful in the Mapping of the Human Brain project. It may also be integrated with functional imaging by PET scan and with the human genome project.

Electrical responses to water‐soluble components of fish mucus recorded from the cnidocytes of a fish predator,Physalia physalis

Physalia physalis, the Portuguese man of war, consumes mostly fish and fish larvae. Intracellular recordings from nematocyst‐containing cells (cnidocytes) in small pieces of Physalia tentacle were used to quantify the electrical responses to diluted and filtered fish skin mucus, 1–100 x 10‐6 M amino acids, monosaccharides, and nucleosides, and seawater, which were delivered upstream of the tissue. Seawater caused responses (one pulse only) in about 10% of the applications. Fish mucus extract elicited responses in all applications, producing 1–18 depolarizing pulses (20 mV maximum amplitude). The pulses were characteristic of post‐synaptic potentials (EPSPs). Lucifer yellow and biocytin dye injections showed that the cnidocytes were not electrically coupled. Simultaneous records from two cnidocytes following mucus applications were identical.We propose, therefore, that the chemoreceptors are not on the cnidocytes, but are probably on sensory neurons that innervate clusters of cnidocytes. A < 3,000 MW fract...

The organization of the turtle inner retina. I. ON- and OFF-center pathways.

Intracellular recordings and dye injections of Lucifer yellow, horseradish peroxidase, or Neurobiotin were made in bipolar, amacrine, and ganglion cells of the Pseudemys turtle retina. By using a standard light-stimulation protocol in a sample of 375 labeled neurons, we were able to identify morphological and physiological characteristics of 11 types of bipolar cell, 37 types of amacrine cell, and 24 types of ganglion cell. To make sense of these data, we have chosen to group the 72 essentially different neuron types into traditional, functionally significant pathways. In this paper we look at the neuronal types in the inner plexiform layer (IPL) in terms of their contribution to generalized luminosity responses such as sustained ON- or OFF-center and transient ON-OFF ganglion cells; in the companion paper (J. Ammermüller, J.F. Muller, and H. Kolb, 1995, J. Comp. Neurol. 358:35-62) we look at them in terms of their involvement in color opponency and directional selectivity. A functional organization of the turtle IPL into OFF sublaminae (strata 1 and 2) and ON sublaminae (strata 3, 4, and 5), as has been described for other vertebrate retinas, was quite clear for two varieties of OFF-center bipolar cells (B4 and B5) and for all four types of sustained ON-center bipolar cell (B1, B2, B6, and B7). Thus, we found no sustained ON-center bipolar cell terminating in strata 1 and 2. We did, however, see three varieties of sustained OFF-center bipolar cells (B3, B9, and B10) having axon terminals in strata 3-5 (the ON sublamina) in addition to their terminations in stratum 1 or 2 (the OFF sublamina). Monostratified sustained ON- and OFF-center amacrine and ganglion cells rigidly obeyed the border of ON and OFF sublaminae. However, multistratified and diffuse sustained amacrine and ganglion cells could be either ON-center or OFF-center, and they did not strictly obey the border: such ON-center cells always had processes in one of the ON sublaminae (strata 3-5), and the equivalent OFF-center cells always had processes in one of the OFF sublaminae (strata 1 and 2). Monostratified transient amacrine and ganglion cells were concentrated in the middle of the IPL (around stratum 3), whereas bi-, tri-, or multistratified transient amacrine or ganglion cells always had processes in both the ON and the OFF sublaminae.(ABSTRACT TRUNCATED AT 400 WORDS)

Expression of a dominant negative inhibitor of intercellular communication in the early Xenopus embryo causes delamination and extrusion of cells

A chimeric construct, termed 3243H7, composed of fused portions of the rat gap junction proteins connexin32 (Cx32) and connexin43 (Cx43) has been shown to have selective dominant inhibitory activity when tested in the Xenopus oocyte pair system. Co-injection of mRNA coding for 3243H7 together with mRNAs coding for Cx32 or Cx43 completely blocked the development of channel conductances, while the construct was ineffective at blocking intercellular channel assembly when coinjected with rat connexin37 (Cx37). Injection of 3243H7 into the right anterodorsal blastomere of 8-cell-stage Xenopus embryos resulted in disadhesion and delamination of the resultant clone of cells evident by embryonic stage 8; a substantial number, although not all, of the progeny of the injected cell were eliminated from the embryo by stage 12. A second construct, 3243H8, differing from 3243H7 in the relative position of the middle splice, had no dominant negative activity in the oocyte pair assay, nor any detectable effects on Xenopus development, even when injected at four-fold higher concentrations. The 3243H7-induced embryonic defects could be rescued by coinjection of Cx37 with 3243H7. A blastomere reaggregation assay was used to demonstrate that a depression of dye-transfer could be detected in 3243H7-injected cells as early as stage 7; Lucifer yellow injections into single cells also demonstrated that injection of 3243H7 resulted in a block of intercellular communication. These experiments indicate that maintenance of embryonic cell adhesion with concomitant positional information requires gap junction-mediated intercellular communication.

Acetylcholine-induced retraction of an identified axon in the developing leech embryo

At early stages of embryonic development of the glossiphoniid leech, Theromyzon rude, a branch, termed MAC, of the axon of the segmentally iterated Retzius neuron extends into the anterior interganglionic connective nerve. At later stages, this branch disappears again in about 30% of the Retzius neurons in the standard midbody segments and in about 75% of the Retzius neurons in the two reproductive midbody segments. The frequency of disappearance of the MAC branch increases to about 85% in all Retzius neurons upon exposure of the embryos to culture media containing 1 mM acetylcholine (ACh) and 10 microM physostigmine during a sensitive period of axon outgrowth. This disappearance represents a retraction of the MAC branch to its point of origin, while other axon branches of the Retzius neuron remained unaffected. In later development, the retracted (medial) MAC branch was replaced by a new (lateral) branch termed LAC. The observations were made using confocal microscopy of fixed embryos stained with anti-5-HT antibody and confirmed by Lucifer yellow injection of individual Retzius neurons. The specific retraction of a single axon branch might be attributable to the local presence of extracellular matrix molecules in the ganglionic neuropil, which is contacted only by the MAC axon branch and could render this branch susceptible to growth-regulating signals. Since Retzius axon morphology in standard segments of ACh-treated embryos resembled that of reproductive segments in untreated embryos, it appears possible that ACh treatment may have simulated a process that contributes to the segmental differentiation of the Retzius neuron.

Targeting and activity-related dendritic modification in mammalian retinal ganglion cells

We have studied factors that influence the development of dendritic morphology in hamster retinal ganglion cells. By combining fluorescent retrograde tracing with in vitro Lucifer yellow injection into fixed retina, cells with appropriate and inappropriate visuotopic projections have been compared. In adult hamsters, cells with an aberrant ipsilateral projection from the nasal retina display a uniformly sparse dendritic morphology. However, following monocular enucleation at postnatal day 0 (P0), this population displays a significantly enhanced dendritic complexity in the adult. By contrast, removal of one eye at P6 or at P12 produces progressively less effect. These results suggest that dendritic complement of the adult aberrant projection can be regulated by altering the early postnatal axonal environment. The development of aberrant ganglion cells was investigated to determine the relative influences of cell death and dendritic remodeling in shaping the composition of the adult aberrant population. Aberrant cells were found to be indistinguishable from other cells in nasal retina throughout early development. After ganglion cell death (P1-P12) is over, aberrant cells still display a full range of cell types. However, at eye opening (P16) they undergo a rapid loss of dendritic complexity by remodeling. By P22, aberrant cells display a uniformly sparse dendritic morphology. When hamsters were raised in the dark between P12 (the end of ganglion cell death) and P22, this severe remodeling was blocked. This block was maintained when hamsters were dark reared to P42. Hence, both dark rearing and monocular enucleation at P0 produce similar effects on the development of visuotopically inappropriate hamster retinal ganglion cells. We speculate that the patterns of dendritic sculpting that we have observed reflect activity-mediated modulation of dendritic form via retrograde signals from the terminal arbors. This has implications for retinal ganglion cell morphological classification and, more generally, for mechanisms that influence the dendritic development of other neurons in the CNS.

Modulation of dorsal thalamic cell activity by the ventral pallidum: its role in the regulation of thalamocortical activity by the basal ganglia.

The actions mediated by limbic system output projections of the basal ganglia were investigated by studying the effects of ventral pallidum (VP) stimulation on the activity of neurons in thalamic target nuclei, including several of the dorsal thalamic nuclei and the nucleus reticularis, using in vivo intracellular recordings in rats. Intracellular injection of Lucifer yellow was used in a subset of experiments to identify the neurons recorded and to confirm their location with respect to the specific thalamic nuclei targeted. Stimulation of the VP evoked ipsps in 79% of the mediodorsal cells recorded. In the reticular nucleus, 73% of the neurons tested responded with evoked ipsps. In contrast, in other dorsal thalamic nuclei VP stimulation evoked depolarizations in 58% of the cells recorded. The latency to onset of the ipsps in the mediodorsal nucleus and in the reticular nucleus were not substantially different (1.7 +/- 1.1 msec vs. 2.7 +/- 1.1 msec), whereas the depolarizing response evoked in dorsal thalamic nucleus neurons typically occurred at longer and more variable latencies (3.5 +/- 2.7 msec). These experiments support a dual functional role for limbic system output from the basal ganglia in the regulation of thalamocortical activity: a) a direct inhibitory projection from the VP to the mediodorsal nucleus and b) an indirect disinhibition of neurons in other dorsal thalamic nuclei that occurs via a direct inhibitory projection to the reticular nucleus of the thalamus. Such an anatomical arrangement may be relevant to the presence of hypofrontality and the breakdown of cognitive filtering observed in schizophrenics.

Intracellular dye injection of previously immunolabeled insect neurons in fixed brain slices

Our method combines intracellular dye injection and immunohistochemistry. Under optical control, Lucifer Yellow was injected into immunohistochemically identified neurons that reside in fixed tissue. The technique allows visualization of the complete arborization patterns of immunostained neurons. Injections were performed on small neurons (somata < 10 μm in diameter). The technique works on microslices of insect brain. Standard immunohistochemical procedures have only been varied slightly, omitting Triton X-100 treatment. Anti-Lucifer Yellow immunohistochemistry, or alternatively the photoconversion technique, enables extension of the morphological analysis of these cells to the electron microscopic level. In the present study, Lucifer Yellow injections were performed on immunohistochemically pretreated brain microslices (anti-Locusta tachykinin II antiserum) of the beetle Tenebrio molitor.

Inverted Pyramidal Neurons and Interneurons in Cat Cortical Subplate Zone are Labelled by Monoclonal Antibody SP1

During development, the subplate zone of the cat neocortex contains neuronal populations with distinct morphological and neurochemical phenotypes. A subset of those are specifically recognized by a mouse monoclonal antibody termed SUBPLATE-1 (SP1), which was generated against tissue homogenates of kitten cortical white matter. SP1 stains cell bodies and proximal dendrites, but rarely distal dendrites, axonal arbors or spines. In order to characterize morphologically the SP1 immunoreactive subplate cell types, we combined SP1 immunohistochemistry with intracellular iontophoretic injections of Lucifer yellow. The majority of double-labelled neurons were inverted pyramids with a single thicker spine-covered dendrite that descended into the white matter and a tuft of thinner spinous dendrites that ascended from the upper somatic pole, but generally remained confined to the white matter. Other double-labelled neurons were multipolar to bitufted, although often equipped with one thicker descending dendrite. In inverted pyramidal cells, the axons originated from the descending dendrite or, more rarely, from the lower portion of the soma, and descended into the white matter. They formed collaterals recurring toward the grey matter. The presence of dendritic spines on double-labelled pyramidal cells and the axonal arborization patterns were two novel features not revealed previously by SP1 immunohistochemistry alone. The inverted pyramidal morphology was typical for double-labelled neurons located in the subplate zone below the apices of the gyri, whereas those located below the flanks or sulci or deep in the white matter often displayed a bitufted or multipolar spinous morphology. A minority of the double-labelled neurons were multipolar with smooth dendrites and locally branching axons. These results suggest that in the cat subplate zone, a majority of the cells expressing the SP1 antigen are spinous, and we termed the spinous subplate cells 'subplate pyramidal neurons'.

Incidence of neuronal coupling in supraoptic nuclei of virgin and lactating rats: estimation by neurobiotin and Lucifer yellow

Dye coupling among neurons has been shown to reflect electrotonic coupling. Recent work in retina has revealed that the incidence of coupling is greater when estimated by neurobiotin (NB) transfer than by Lucifer yellow (LY). Several previous studies have shown that the incidence of LY coupling among supraoptic nucleus (SON) neurons of lactating rats is 2- to 4-fold higher than is observed in virgin females. We compared the incidence of coupling among SON neurons following simultaneous injections of LY and NB into the same cells in slices from virgin or lactating rats. As seen in previous studies, there were 4-fold more LY-coupled neurons per injection in lactating as compared to virgin rats. Under both conditions, the numbers of NB-coupled neurons per injection were 4-fold greater than was observed for LY; possible mechanisms are discussed. Individual NB-filled neurons were coupled to as many as 10 other cells distributed over a large area of the SON. These results confirm previous findings of more coupling in lactating than virgin SONs, and suggest that both the incidence and spatial extent of interneuronal coupling are greater and thus probably more important functionally than has been heretofore appreciated.

Developmental regulation of voltage-gated K+ channel and GABAA receptor expression in Bergmann glial cells

Bergmann glial cells are closely associated with neurons: during development they provide guiding structures for migrating granule cells and in the adult cerebellum they display intimate interactions with Purkinje cells. In this study, we have addressed the question of whether such changes in neuronal-glial interactions during development are accompanied by variations in the membrane properties of Bergmann glial cells. We used a mouse cerebellum slice preparation to study membrane currents of the Bergmann glial cells at various stages of development in situ using the patch-clamp technique. The distinct morphology of Bergmann glial cells was revealed by Lucifer yellow injections during recording. While Bergmann glial cells in mice of postnatal day 20 (P20) to P30 have thick processes with arborized, irregularly shaped leaf-like appendages, the processes of cells from younger mice (P5-P7) are thinner and smoother. This morphological maturation is accompanied by a variation in voltage-gated currents. In cells from P5 to P7, delayed outward- and inward-rectifying K+ currents were recorded, while older Bergmann glial cells were characterized by, large, voltage- and time-independent K+ currents. In addition, application of GABA induces two effects, a rapid activation of a Cl- conductance and a longer-lasting decrease in the (resting) K+ conductance. Both effects were mediated by benzodiazepine-insensitive GABAA receptors. Responses in cells of P5-P7 mice were large as compared to the small or even undetectable responses in P20-P30 cells. These GABAA receptors were characterized immunohistochemically in mice and rat brain sections with five subunit-specific antibodies. Bergmann glial cells exhibit a distinct but transient immunoreactivity for the GABAA receptor alpha 2-, alpha 3-, and delta-subunits. Staining is maximal between P7 and P10 and decreases gradually thereafter. In contrast, antibodies to the alpha 1- and beta 2,3-subunits fail to decorate Bergmann glial cells, although they yield a prominent staining of both the Purkinje cells and the granule cells. These changes in the Bergmann glial cell membrane properties and GABAA receptor expression suggest a transition between functional states during development of the Bergmann glial cells.

Intracellular characterization of identified sensory cells in a new spider mechanoreceptor preparation.

1. We have developed an isolated mechanoreceptor-organ preparation in which the intact sensory structures are available for mechanical stimulation and electrical recording. The anterior lyriform slit sense organ on the patella of the spider, Cupiennius salei Keys., consists of seven or eight cuticular slits, each innervated by a pair of large bipolar sensory neurons. The neurons are fusiform, and the largest somata are < or = 120 microns long. The innervation of the organ was characterized by light microscopy of neurons backfilled with neuronal tracers. Intracellular recording was used to measure the passive and active electrical properties of the neurons, in several cases followed by identification with Lucifer yellow injection. Both neurons of each pair from one slit responded with action potentials to depolarization by a step current injection. Approximately half of the sensory neurons adapted very rapidly and generated only one or two action potentials in response to a sustained depolarizing step, while a second group produced a burst of action potentials that adapted to silence in approximately 1 s or less. Recordings from identified neuron pairs indicated that each pair consists of one rapidly adapting and one bursting neuron. Measurements of cell membrane impedances and time constants produced estimates of neuronal size that agreed with the morphological measurements. This new preparation offers the possibility of characterizing the mechanisms underlying transduction and adaptation in primary mechanosensory neurons.

Morphological assessment of grafted rat and mouse cortical neurons: A light and electron microscopic study

The morphology of cortical neurons grafted into (or near) the rat striatum was studied by means of intracellular Lucifer yellow injections in fixed slices. Rat donor syngeneic cortical tissue (from postnatal day 1 old rats; AO strain) as well as mouse donor xenogeneic cortical tissue (prenatal day 19; C3H/HE strain) were grafted as solid pieces into 8-12 week-old rats (AO strain). Recipients of mouse xenografts were immunosuppressed with a monoclonal antibody against the interleukin-2 receptor. After perfusion and sectioning of the graft-containing areas, individual slices were incubated in the DNA stain 4.6-diamidino-2-phenylindole (DAPI) to visualize the cell nuclei. Grafts could be easily identified by a surrounding rim of astrocytes which outline the border between grafted and host tissue. Grafted cortical neurons were intracellularly filled with Lucifer yellow, DAB-photoconverted, and further processed for light and electron microscopy. In general, no cortical lamination could be observed in the grafted rat and mouse cortical tissue, but neurons were loosely packed throughout the graft. Two major cell types could be identified in all grafts investigated so far. The majority resembled those described as spiny neurons (85%), which could be further classified into pyramid-like, spiny stellate-like or fusiform spiny neurons, with somata ranging between 15 and 25 microns in diameter. The remaining 15% resembled non-spiny neurons with either a multipolar basket-like or fusiform morphology. Dendrites of spiny and non-spiny neurons, which could extend to distances up to 400 microns, were never seen to cross the astrocytic border, but some main axon and axonal collaterals of spiny neurons were found to leave the graft. On the basis of light microscopic observations no difference was found between mouse and rat grafted cortical neurons. The results of this study show that grafted cortical neurons retain some of the characteristic features of neurons in the intact adult cerebral cortex, although there appears to be a greater preponderance of spiny neurons in grafted tissue. This may reflect an immaturity of the grafted tissue or a response to the striatal environment.

Unexpected presence of neurofilaments in axon-bearing horizontal cells of the mammalian retina

In several mammals only one of the two types of retinal horizontal cell, the axonless A-type, appears to express neurofilaments. Neurofilament immunostaining of rodent retinas reveals a horizontal cell plexus that has previously been interpreted as belonging to A-type cells. Our intracellular Lucifer yellow injections strongly suggest that there are no A-type horizontal cells in rat and gerbil. Counterstaining of dye-injected cellular structures with a neurofilament antibody directly shows that the axon terminal systems of the axon-bearing B-type horizontal cells contain neurofilaments. These unexpected findings explain and reinterpret the neurofilament plexus in rodent retinas. In contrast, Lucifer yellow injections in guinea pig retina reveal both A- and B-type horizontal cells, showing that horizontal cell types are not uniform among rodents. In the guinea pig retina both A-type cells and B-type axon terminal systems contain neurofilaments.