Intracellular Injection Of HRP Research Articles

Axons of sacral preganglionic neurons in the cat: II. Axon collaterals.

Axon collaterals were identified in 21 of 24 preganglionic neurons in the lateral band of the sacral parasympathetic nucleus of the cat. Following the intracellular injection of HRP or neurobiotin the axons from 20 of these neurons were followed and 53 primary axon collaterals were found to originate from unmyelinated segments and from nodes of Ranvier. Detailed mapping done in the five best labeled cells showed bilateral axon collaterals distributions up to 25,000 microm in length with 950 varicosities and unilateral distributions up to 12,561 microm with 491 varicosities. The axon collaterals appeared to be unmyelinated, which was confirmed at EM, and were small in diameter (average 0.3 microm). Varicosities were located mostly in laminae I, V, VII, VIII and X and in the lateral funiculi. Most varicosities were not in contact with visible structures but some were seen in close apposition to Nissl stained somata and proximal dendrites. Varicosities had average minor diameters of 1.3 microm and major diameters of 2.3 microm. Most were boutons en passant while 10-20% were boutons termineaux. EM revealed axodendritic and axoaxonic synapses formed by varicosities and by the axons between varicosities. It is estimated that the most extensive of these axon collaterals systems may contact over 200 spinal neurons in multiple locations. These data lead to the conclusion that sacral preganglionic neurons have multiple functions within the spinal cord in addition to serving their target organ. As most preganglionic neurons in this location innervate the urinary bladder, it is possible that bladder preganglionic neurons have multiple functions.

Cell fate analysis of teloblasts in the Tubifex embryo by intracellular injection of HRP.

As in other clitellate annelids, embryonic development in the oligochaete Tubifex is characterized by the generation of five bilateral pairs of teloblasts (designated M, N, O, P and Q), which serve as embryonic stem cells to produce germ bands on either side of the embryo. A large part of the tissues comprising body segments has been assigned to the progenies of the teloblasts; however, the developmental fate of each teloblast has been inferred only from its initial position in the embryo. In the present study, the fate of the progenies of each teloblast was followed by means of intracellular injection of a tracer enzyme, horseradish peroxidase. Cell fate maps for teloblasts in the Tubifex embryo were constructed. M teloblasts gave rise to nearly all of the mesodermal tissues, which included circular and longitudinal muscles, coelomic walls, nephridia (in segments VII and VIII) and primordial germ cells (in segments X and XI). Although few in number, M teloblasts also contributed cells to the ventral ganglion. Similarly, each of the ectoteloblasts, N, O, P and Q, made a topographically characteristic contribution to the ectodermal tissues such as the nervous system (i.e. ganglionic cells and peripheral neurones) and epidermis, all of which exhibited a segmentally repeated distribution pattern. The P and Q teloblasts uniquely gave rise to additional ectodermal tissues, namely ventral and dorsal setal sacs, respectively. Furthermore, O teloblasts made a contribution to the nephridiopores in segments VII and VIII as well. These results confirm the previously held view that ectoteloblasts and mesoteloblasts are the main source of ectodermal and mesodermal segmental tissues, respectively, but also suggest that all of the teloblasts produce more types of tissue than has previously been thought.

Synaptic connections between GABA-immunoreactive neurons and uniglomerular projection neurons within the antennal lobe of the cockroach, Periplaneta americana.

Synapses within deutocerebral glomeruli between GABA-immunoreactive, putatively inhibitory local interneurons and uniglomerular projection (output) neurons were demonstrated by means of a combination of GABA-immunogold labeling and intracellular HRP injection. The following connections were identified. 1) GABA-immunoreactive (GABAir) neurons form output synapses in a dyadic fashion onto a uniglomerular projection neuron and, in addition, a second GABAir neuron. A uniglomerular projection neuron in turn forms dyadic output synapses onto two GABAir neurons. Several examples of reciprocal connections have been identified between, first, GABAir neurons and uniglomerular projection neurons, and, second, GABAir neurons themselves. 2) GABAir neurons are serially connected with uniglomerular projection neurons via interposed GABAir processes. In some cases, also the first GABAir process of such a polysynaptic connection formed an output synapse onto the projection neuron. Such serial connections may form the structural basis for both, the feedforward inhibition as well as the feedforward disinhibition of uniglomerular projection neurons by GABAergic neurons. The reciprocal contacts may serve as control devices that modulate the output activity of the projection neurons.

Morphology, dendritic field size, somal size, density, and coverage of M and P retinal ganglion cells of dichromatic Cebus monkeys

Male Cebus monkeys are all dichromats, but about two thirds of the females are trichromats. M and P retinal ganglion cells were studied in the male Cebus monkey to investigate the relationship of their morphology to retinal eccentricity. Retinal ganglion cells were retrogradely labeled after optic nerve deposits of biocytin to reveal their entire dendritic tree. Cebus M and P ganglion cell morphology revealed by biocytin retrograde filling is similar to that described for macaque and human M and P ganglion cells obtained by in vitro intracellular injection of HRP and neurobiotin. We measured 264 and 441 M and P ganglion cells, respectively. M ganglion cells have larger dendritic field and cell body size than P ganglion cells at any comparable temporal or nasal eccentricity. Dendritic trees of both M and P ganglion cells are smaller in the nasal than in the temporal region at eccentricities greater than 5 mm and 2 mm for M and P ganglion cells, respectively. The depth of terminal dendrites allows identification of both inner and outer subclasses of M and P ganglion cells. The difference in dendritic tree size between inner and outer cells is small or absent. Comparison between Cebus and Macaca shows that M and P ganglion cells have similar sizes in the central retinal region. The results support the view that M and P pathways are similarly organized in diurnal dichromat and trichromat primates.

Quantitative ultrastructure of Ia boutons in the ventral horn: scaling and positional relationships

Three physiologically characterized spindle (group Ia) afferents were labeled by the intracellular injection of HRP and were processed for light-level reconstruction. Thirty-five boutons in the ventral horn were then selected for analysis. They were serially thin sectioned and characterized in terms of volume, total surface area and the surface area of apposition to postsynaptic neurons (apposed surface area), mitochondrial volume, vesicle and active zone features, relation to presynaptic contacts, postsynaptic profile size, and position within the terminal arbor. Virtually all of these characteristics were widely variable, both within the entire population and in the endings of a single fiber. Apposed surface area, mitochondrial volume, vesicle number, active zone vesicle number, active zone number, and total active zone area were highly correlated in a positive linear manner with bouton volume. This suggests a type of ultrastructural "size principle," in which the morphological features associated with synaptic release scale directly in proportion to bouton size. This pattern also extends to local circuit interactions: the extent of an Ia bouton's input from axoaxonal contacts (86% receive at least one axoaxonal contact) was directly proportional to its size. In addition, the characteristics of an Ia bouton were related to its position on the postsynaptic element and within the terminal arbor. Vesicle density, percentage mitochondrial volume, and active zone size increased as the postsynaptic process decreased in size, while volume, apposed surface area, active zone number and area, and vesicle number all decreased as one moved downstream within a terminal branch, with the exception of the terminal bouton. Vesicle density also decreased as one moved away from the dorsal root entry zone.

Correlations between the receptive field properties and morphology of neurons in the deep layers of the hamster's superior colliculus

Extracellular and intracellular recording, receptive field mapping, and intracellular HRP injection techniques were used to define the morphological classes of cells in the deep laminae of the hamster's superior colliculus and to determine whether there are any correlations between the structural and functional characteristics of these neurons. A total of 110 neurons were characterized and reconstructed. Of these, 23.6% (N = 26) were visual, 60% (N = 66) were somatosensory, 0.9% (N = 1) were bimodal (visual-somatosensory), and 15.4% (N = 17) were unresponsive. Of the somatosensory neurons, 72.7% (N = 48) were low threshold, 4.5% (N = 3) had a wide dynamic range, 9.1% (N = 6) responded only to noxious stimulation, and 13.6% (N = 9) had complex somatosensory receptive fields. Deep layer cells were divided into eight morphological classes. These classes were multipolar cells (26.4%, N = 29), bipolar cells (9.1%, N = 10), widefield vertical cells (7.3%, N = 8), horizontal cells (13.6%, N = 15), stellate cells (10.9%, N = 12), ventrally directed cells (5.5%, N = 6), sparse radial cells (17.3%, N = 19), and small sparse radial cells (6.4%, N = 7). Four cells (3.6%) did not fit into this classification scheme. Univariate and multivariate analyses of variance of properties such as soma area, number of branch points, total dendritic length, and volume and orientation of dendritic arbor indicated that these classes were significantly different. However, chi 2 analysis and multivariate analysis of variance indicated no significant relationships between morphological class and either laminar location or receptive field type. There was a significant positive relationship between the possession of dendrites that extended into the superficial laminae and visual responsivity.

A parsimonious description of motoneuron dendritic morphology using computer simulation

Most quantitative descriptions of neuronal dendrite morphology involve tabulations of measurements and correlations among them. The present work is an attempt to extract from such data a parsimonious set of parameters that are sufficient to describe the quantitative features of individual and pooled dendrites, including their statistical variability. A relatively simple stochastic (Monte Carlo) model was devised to simulate branching dendritic trees. The necessary parameters were then derived directly from measurements of 64 completely reconstructed dendrites belonging to six gastrocnemius alpha-motoneurons, labeled by intracellular injection of HRP. Comparison of actual and simulated dendrites was used to guide the process of parameter extraction. The model included only two processes, one to generate individual branches given their starting diameters and the second to select starting diameters for the daughter branches produced at dichotomous branching points. The stochastic process for branch generation was controlled by probability functions for branching (Pbr) and for terminating (Ptrm), together with a constant rate of branch taper. All model parameters were fixed by motoneuron measurements except for branch taper rate, which was allowed to vary within limits consistent with observed taper rates in order to generate the appropriate total number of branches. The simplest model (model 1), in which Pbr and Ptrm depended only on local branch diameter, produced simulated dendrites that fit many, but not all, characteristics of actual motoneuron dendrites. Two additional properties produced significant improvements in the fit: (1) a small but significant dependence of daughter diameters on the normalized starting diameter of the parent branch, and (2) a dependence of Pbr and Ptrm on distance from the soma as well as on local branch diameter. The process of developing this model revealed unsuspected relations in the original data that suggest the existence of fundamental mechanisms for morphological control. The final model succinctly describes a large amount of data and will enable quantitative comparisons between the dendritic structures of different types of neurons, regardless of their relative sizes.

Functional influence of interlaminar connections in the hamster's superior colliculus

The aim of this study was to determine the functional importance of intrinsic connections within the hamster's superior colliculus (SC) in the development of the visual responses of neurons in the deep layers of this nucleus. We used localized injections of lidocaine to block action potentials of fibers that passed through the superficial layers and injections of CoCl2 to block synaptic transmission in these layers. We also determined the morphology of a number of the deep layer cells recorded in these experiments by intracellular injection of HRP. Injection of lidocaine into the superficial layers completely abolished the visual- and/or optic chiasm-evoked responses of all 40 deep layer cells tested. Thus, fibers that either pass through or synapse in the superficial layers are necessary for the visual responses of deep layer neurons. Injections of CoCl2 restricted to the superficial layers significantly reduced the visual responsivity of 86% of 92 deep layer neurons tested and abolished the visual responses of 68% of these cells. Superficial layer injections of CoCl2 were equally effective in reducing the responses of neurons with dendrites that ascended into the superficial layers (all seven cells tested and recovered) and those of cells with dendrites restricted to the deep layers (six of seven cells tested and recovered). Injections of CoCl2 into the deep layers, in the region of the cell being recorded, significantly reduced the visual responses of 59% of 37 cells and abolished the visual responses of 40% of the neurons tested. Deep CoCl2 injections abolished the visual responses of three of four cells with dendrites restricted to the deep layers and only one of four cells with dendrites that ascended into the superficial layers. These results indicate that descending interlaminar axons from visually responsive superficial layer cells may be responsible for the visual responsivity of most neurons in the deep SC layers of hamster. Also, some deep layer neurons in this species may receive effective visual input through their dendrites that ascend into the superficial layers, where they are likely to be contacted by retinal axons or axon collaterals of superficial layer cells.

Intracellular HRP study of nociceptive neurons within the ventrobasal complex of the cat thalamus

Locations and morphological characteristics of nociceptive specific (NS) and wide dynamic range (WDR) neurons of the ventrobasal (VB) thalamic complex were studied using an intracellular HRP injection technique in cats anesthetized with urethane and chloralose. Both NS and WDR neurons were found within the marginal zone of the VB complex. Their somata were confined to the VB complex, but dendrites were extended into the structures of the thalamus surrounding the VB complex. NS neurons were found in the caudal part of the VB complex, whereas WDR neurons were found more rostrally. Examinations of their locations ithin the marginal zone of the VB complex confirmed the somatotopic organization, as found previously. We also confirmed the previous reports that there are 3 different types of low-threshold mechanoreceptive (LTM) neurons within the VB complex. They are type I, type II and type III neurons. Both NS and WDR neurons were either type I or type II neurons. Obvious morphological differences were not found between LTM neurons and two classes of nociceptive VB neurons investigated.

Axonal regeneration of retinal ganglion cells in the cat geniculocortical pathway

The optic nerve of anesthetized cats was completely cut and the autologous sciatic nerve was transplanted. Sixty days later some populations of retinal ganglion cells were shown to regenerate the axon with retrograde HRP labeling. We verified that ganglion cells that had projected to the lateral geniculate nucleus (LGN) were able to regenerate through the transplant with a double-labeling method: diI was injected into the LGN prior to the transplantation, and dextran-fluorescein was injected into the graft after axonal regeneration. Intracellular injection of HRP into regenerating ganglion cells in an in vitro preparation revealed that the two major cell types projecting to the LGN, α and β, regenerated axons and showed normal dendritic morphology.

Trigeminal nociceptive neurons in the subnucleus reticularis ventralis. I. Response properties and afferent connections

Trigeminal nociceptive neurons within the subnucleus reticularis ventralis medullae oblongatae (SRV), which lies ventral to the trigeminal subnucleus caudalis and subnucleus reticularis dorsalis medullae oblongatae, were studied in urethane/chloralose-anesthetized cats and monkeys. These neurons were called ‘SRV neurons’. They were almost regularly excited by pressure to the ipsilateral cornea or to both corneas at a strength well above the human corneal pain threshold. Most of them were activated by noxious mechanical stimulation of the pinna, face and/or tongue. A significant fraction of SRV units was responsive to tapping of the ipsilateral dorsum of the nose and/or electrical stimulation of tooth pulp afferents. Evidence was obtained that responses to tapping of the dorsum of the nose were due to mechanical distortion of the nasal mucosa. Intracellular injection of HRP into SRV neurons demonstrated that injected neurons were large neurons characteristic of the SRV. Trigeminal tractotomy just rostral to the obex did not eliminate responses of SRV units to trigeminal inputs. Neurons relaying trigeminal inputs to SRV neurons were electrophysiologically identified in the nucleus reticularis parvocellularis which is ventromedially adjacent to the subnuclei oralis and interpolaris of the trigeminal spinal tract nucleus. These findings were supported by HRP injection into the SRV. Units having receptive fields similar to those of SRV neurons were found in lamina VII of the first cervical cord, suggesting that SRV neurons may be trigeminal lamina VII neurons.

Dendritic morphology of pyramidal neurones of the visual cortex of the rat: I. Branching patterns

The aim of this study was to provide quantitative descriptions of the branching patterns of basal and apical dendrites of pyramidal neurones from the visual cortex of the rat. Thirty-nine neurones from cortical layers 2/3 and 5, that had been injected with horseradish peroxidase, reconstructed, and measured with the light microscope as part of an earlier study (Larkman and Mason, '90; J. Neurosci. 10:1407-1414), were used. The cells had previously been divided into three classes, layer 2/3 cells and thick and slender layer 5 cells, on the basis of their dendritic morphology. The branching pattern of the basal and apical oblique dendrites was similar for all the cells. Between 3 and 9 basal trees arose from the soma and the number of tips in each tree varied widely, between 1 and 13. The path lengths of all the basal dendrites of a given cell were relatively constant, however. Most basal dendritic branching occurred close to the soma, such that terminal segments were much longer than intermediate segments and contributed approximately 90% of the total dendritic length of each tree. Terminal segments showed only a narrow range of diameters. Most apical oblique trees arose from the proximal part of the apical trunk. They tended to be less highly branched but were otherwise extremely similar to basal trees. Distal oblique trees were unbranched or branched only once, and their terminal segments tended to be shorter and thinner than those of basal trees. The branching pattern of the apical terminal arbors was different, with many longer intermediate segments. The terminal segments tended to be thinner than those of basal or proximal oblique trees. Slender layer 5 cells were without obvious terminal arbors. The basal and proximal oblique dendrites jointly sampled a roughly spherical volume of cortex centred about the soma, and together they accounted for the substantial majority of the cell's total dendritic shaft membrane area. Comparisons with previous studies suggest that intracellular HRP injection can yield a more complete visualization of dendritic morphology than is obtained using Golgi-based methods (unless cells are reconstructed across tissue slabs), and can therefore result in a different view of the relative importance of the various components that make up the cell's dendritic system.

The need for cellular elements during axonal regeneration in the sea lamprey spinal cord

Spinal axons in the larval sea lamprey regenerate following a complete spinal transection. It is not known whether regenerating growth cones require contact with cellular elements or whether the basement membrane and collagenous meninx primitiva which surround the spinal cord are sufficient for neurite outgrowth. To determine this, a freeze lesion was made which severed axons, destroyed neuronal perikarya, and greatly reduced the number of glial cells. After at least 10 weeks of recovery, 50 neurites from 31 Müller and Mauthner axons were labeled by intracellular injection of HRP. Eighty-six percent of these neurites did not regenerate into the lesion site. No neurites grew through the lesion. No animals recovered coordinated swimming. These results suggest that glial and/or neuronal surfaces are required for axonal regeneration. Moreover, a monolayer of glial cells appears to be suboptimal and a three-dimensional matrix of cells may be necessary to promote regeneration in the lamprey spinal cord.

Dendritic and axonal morphology of HRP‐injected neurons in the inferior colliculus of the cat

The dendritic and axonal morphology of neurons in the inferior colliculus of the cat was investigated after intracellular injection of HRP, in vivo. All injected axons gave off local collaterals, and most showed a widespread distribution and lacked a specific orientation. In contrast, the dendrites of injected neurons were distinguished by their degree of orientation and the direction of the longest axis of orientation. Dendrites showed a high, moderate, or low degree of orientation. Most highly oriented cells had their longest axis in the rostrocaudal direction with fewer in the mediolateral direction. In the central nucleus, only the rostrocaudally oriented cells correspond to the disc-shaped cells identified in Golgi preparations. Unlike most cells in our sample, the two cells that were disc-shaped had axons that were parallel to the orientation of the dendritic tree. In the dorsal cortex, rostrocaudally oriented cells also were found, but they had unoriented axons. In both the central nucleus and dorsal cortex, cells with a mediolateral axis of orientation or no specific orientation correspond to stellate cells and had axons with widespread local collaterals. These results suggest that an extensive network of local axon collaterals may contribute to neural processing within the inferior colliculus. In the central nucleus, local axons may establish connections within or across the fibrodendritic laminae. In the dorsal cortex, the local and afferent axons may form a complex reticular network. Finally, some injected cells had axons terminating locally and also entering the brachium of the inferior colliculus. This suggests that cells in the inferior colliculus may function as both interneurons and projection neurons.

Correlations between morphology and electrophysiology of pyramidal neurons in slices of rat visual cortex. I. Establishment of cell classes

The purpose of this study was to determine whether different classes of neocortical pyramidal neurons possess distinctive electrophysiological properties. Neurons were assigned to classes on the basis of their somadendritic morphology as revealed by intracellular HRP injection. The electrophysiological features of each class are compared in the following paper (Mason and Larkman, 1990). Coronal slices were prepared from the visual cortex of the albino rat and maintained in vitro. Pyramidal neurons from layers 2/3 and 5 were impaled with electrodes containing HRP. After intracellular recording, the cells were injected, reacted to visualize the enzyme, and examined in the light microscope. The 18 neurons from layer 2/3 showed an underlying similarity of somadendritic morphology and were considered to constitute a single cell class, although some features varied with the depth of the soma in the cortex. The 22 layer 5 cells were more diverse and were divided into 2 main classes. Eleven cells had single, thick apical trunks which ascended to close to the layer 1 border then branched to form a terminal arbor. These cells were termed thick layer 5 cells, and were found mainly in the upper part of the layer. Ten layer 5 cells had single apical trunks which tapered to a fine diameter and terminated, without forming an obvious terminal arbor, near the top of layer 4 (3 cells) or in layer 2/3 (7 cells). These cells were termed slender layer 5 cells and were found throughout the layer.(ABSTRACT TRUNCATED AT 250 WORDS)

Physiological and morphological characteristics of cat masticatory motoneurons—intracellular injection of HRP

The physiology and morphology of masticatory motoneurons of adult cats were examined by the methods of intracellular recording and intracellular injection of horseradish peroxidase. Masseter and jaw-opening motoneurons were identified by intracellular recordings of the antidromic response following stimulation of the masseter and mylohyoid nerves, respectively. An excitatory postsynaptic potential (EPSP) was recorded from masseter neurons by stimulation of the masseter nerve with stimulus intensity below threshold for antidromic response. In contrast, the EPSP was not recorded from jaw-opening motoneurons by stimulation of the mylohyoid nerve with stimulus intensity below threshold for antidromic response. Patterns of postsynaptic potentials (PSPs) in the masseter motoneurons following stimulation of the tooth pulp or periodontal afferents were classified into 4 types: hyperpolarization ( n = 40), depolarization-hyperpolarization ( n = 9), hyperpolarization-depolarization ( n = 5), and depolarization with spike potentials ( n = 10). On the other hand, patterns of the PSPs in the jaw-opening motoneurons following stimulation of the same afferents were classified into two types: depolarization with spike potentials ( n = 19), and hyperpolarization ( n = 5). Twenty-five masseter and 7 jaw-opening motoneurons and an intranuclear neuron were reconstructed from serial sections in the transverse plane. On the basis of dendritic morphology, the masseter motoneurons could be classified into two major groups, type I ( n = 15) and type II ( n = 9), whereas two neurons were found to constitute a separate category of the masseter motoneuron. The dendritic distributions of all the jaw-opening motoneurons examined were generally similar and there was no indication of the existence of subtypes, whereas there were 2 or 3 subgroups in type I and type II masseter motoneurons. Type I masseter neurons had primary dendrites which extended radially in all directions, and the whole profile of their dendritic trees presented a spherical and an egg-shaped appearance. In type II masseter neurons, the origin of primary dendrites was bipolar or tripolar, and the whole profile of their dendritic trees presented a hemispherical and mirror-imaged, funnel-shaped appearance. The other two masseter motoneurons had a particular dendritic tree which was much simpler in configuration, with less tapering or branching than those of other neurons examined. In contrast, the dendritic profiles of all the jaw-opening motoneurons were similarly organized and showed vertically oriented dendritic trees which were more developed in the dorsomedial than in the ventrolateral direction. Dendrites from all groups extended beyond the boundaries of the motor nucleus into the reticular formation. The border between the dorsolateral and ventromedial subdivisions was crossed by some dendrites but the overlap was small. Among neurons examined in the present study, one intranuclear neuron identified had the simplest dendritic tree and smalles soma size, with axon collaterals projecting to the reticular formation. No significant differences in several soma-dendritic parameters were found between morphologically distinct motoneuronal subpopulations and between the jaw-opening and masseter motoneurons. There were no instances in which axon collaterals were observed for the masticatory motoneurons.

Morphology and synaptic connections of HRP-filled, axon-bearing horizontal cells in the Xenopus retina.

Axon-bearing horizontal cells of the Xenopus retina were studied by intracellular injection of HRP following physiological characterization. The profile of the cell viewed in whole mount consisted of a round or oval perikaryon about 50 microns in diameter and an axon about 1 mm long which lacked a prominent terminal expansion. The axonal diameter was 0.5-1.0 microns in its proximal third but 2-4 microns in its distal portion. Along its course the axon emitted 25-40 branchlets each 0.2 micron in diameter, up to 10 micron long and terminating in a cluster of two to six synaptic knobs. Electron microscopic examination revealed that both perikaryal dendrites and axon branchlets ended in both rod and cone synaptic bases; cone contacts outnumbered rod contacts by two- to threefold. We were unable to document synapses of presumed interplexiform cells onto identified horizontal cells. Horizontal cell axons are joined in their distal portions by numerous, small (0.2 micron long) gap junctions. Other gap junctions were noted between horizontal cell processes within the synaptic endings of photoreceptors. An hypothesis is advanced whereby the cluster of axon branchlet synaptic knobs permits dynamic interaction of rod and cone synaptic inputs to the horizontal cell.

Morphology of neurons in area 4γ of the cat's cortex studied with intracellular injection of HRP

Neurons in laminae II, III, V, and VI of area 4 gamma of the cat motor cortex were studied following intracellular penetration with an HRP-filled micro-electrode. Antidromic and synaptic responses produced by stimulation of the cerebral peduncles and/or of the ventrolateral nucleus of the thalamus were investigated. Horseradish peroxidase was then iontophoresed into the same neurons to allow examination of their detailed morphology. The morphology of pyramidal neurons whose somata were located in a particular lamina was similar but differed from that of pyramidal neurons in other laminae. The modified pyramidal neurons of lamina II had a truncated apical dendrite or did not possess an obvious apical dendrite, even though the ascending dendritic branches were longer and more extensive than the "basal" branches. As was the case for the pyramidal cells in other laminae, the axons of these lamina II modified pyramidal cells descended toward the white matter; their somata were generally pyramidal in shape, and their dendrites were spiny. All pyramidal neurons except some of lamina VI had ascending dendrites which terminated in a tuft in lamina I, subpially. No intracortical collaterals were seen originating from the axons of lamina II or of lamina VI pyramidal neurons. Lamina III pyramidal neurons had extensive short and long axon collaterals which contributed synaptic boutons to all laminae of the cortex. Pyramidal neurons of lamina V had fewer axon collaterals whose synaptic boutons were restricted to laminae V and VI. All somata of pyramidal tract neurons (PRNs), identified by antidromic responses from peduncular stimulation, were located in lamina V, except for one which was located in lamina VI. Recurrent collaterals of pyramidal neurons were activated by peduncular stimulation. Recurrent excitatory postsynaptic potentials (epsps) could be evoked in fast PTNs, slow PTNs, other pyramidal neurons of lamina V, and pyramidal neurons of lamina VI at latencies between 1.3 and 6.25 msec. In some slow PTNs, a recurrent inhibitory postsynaptic potential of long duration was the predominant response. Stimulation of the ventrolateral nucleus of the thalamus resulted in epsps in pyramidal neurons of lamina III, V, and VI at latencies between 1.0 and 5.0 msec.

Relationship of the axonal and dendritic geometry of spiny projection neurons to the compartmental organization of the neostriatum.

Intracellular injection of HRP combined with immunocytochemistry for [Leu]enkephalin was used to demonstrate striatal spiny neuron dendritic and local axonal arborizations in the same section as enkephalin-rich patches (striosomes). Cobalt intensification of the first DAB reaction prior to the immunoperoxidase steps resulted in good contrast between the black reaction product in the intracellularly labeled cells and the brown staining for [Leu]enkephalin. Serial reconstructions of the labeled cells and nearby boundaries between the enkephalin-rich striosomes and enkephalin-poor matrix allowed the relationship between the arborizations of the labeled cells and these boundaries to be established. It was also possible to examine the relationship to compartmental boundaries of a second neuronal class consisting of large, pallidallike neurons whose somatodendritic morphology was outlined by immunoperoxidase-labeled terminals. We found that spiny projection neurons in both compartments have dendritic arbors and local axonal collaterals that are confined by compartmental boundaries. The termination or recurvature of dendrites at such boundaries suggests that the cellular basis of striatal compartmental organization is provided by this class of striatal neuron. On the other hand, large pallidumlike striatal neurons were found to have dendrites that extend across compartmental boundaries. These results support previous reports that striatal spiny projection neurons preserve the compartmental segregation of parallel striatal input-output systems, whereas other classes of striatal neurons may serve to provide limited integration between compartments.

Giant Aplysia neuron R2 has distal dendrites: evidence for protein sorting and a second spike initiation site

Because of its anatomy, the neuron R2 of Aplysia has been used to study how proteins are distributed to their appropriate destinations within the cell. The R2 cell body resides in the abdominal ganglion, while its axons terminate on glands in the skin. Using intracellular injection of HRP and intraxonal recordings, we found that R2 has a dendritic (receptive) arborization in the pleural ganglion. The structure of these dendrites was examined after injecting the soma with 3H-L-fucose, thereby labeling glycoproteins that are transported to all regions of the cell. Light- and electron-microscope autoradiography show that the openings to the dendrites are not on the periphery, but are suspended inside the axon by glial cell infoldings. All of the organelles seen in the axon are found in the dendrites, including 2 types of vesicles. Neither the axon nor the dendrites contain ribosomes. Thus, R2 has 3 functionally distinct regions--cell body, dendrites, presynaptic terminals--that are separated from each other by at least 4 cm. This implies that pre- and postsynaptic proteins made in the cell body are transported along the axon to the pleural ganglion, where they are sorted. To investigate this idea, we exposed the abdominal ganglion to 35S-methionine to label R2's proteins. Analyses by SDS-PAGE of the rapidly transported labeled proteins from R2 consistently showed a 78 kDa band that accumulated in the pleural ganglion and did not move into the peripheral nerves. This then is a putative dendritic constituent.