Modality-specific axonal projections in the CNS of the flies Phormia and Drosophila

There is a rich history of behavioral and physiological studies on the leg sensory systems of flies. Here we examine the anatomy of the sensory axons of two species of fly and demonstrate that the location of the axonal projections in the CNS can be correlated with the modality they encode. We studied receptors associated with proprioceptive, tactile, and multimodal hairs. Proprioceptive hairs occur in clusters, called hair plates, and are situated near joints. The neuron innervating each proprioceptive hair has a large axon and coarse arborization in the intermediate neuropil. Tactile receptors have smaller arbors, which are located in a ventral region of the thoracic neuromere. Finally, the multimodal hairs are each innervated by one tactile and four chemosensory neurons. The single tactile neuron has a central arbor that is indistinguishable from those of the tactile hairs; the four chemosensory neurons project to yet a third region of neuropil near the ventral surface of each neuromere. Thus there is a clear modality-specific segregation of axonal arbors in the CNS. This organization is identical in Phormia and Drosophila and thus apparently highly conserved within the Diptera. We presume that, as in other insect sensory systems, this anatomical specificity is linked to synaptic specificity.     

Position-specific central projections of mechanosensory neurons on the haltere of the blow fly,Calliphora vicina

The halteres of Dipteran insects play an important role in flight control. They are complex mechanosensory devices equipped with approximately 400 campaniform sensilla, cuticular strain gauges, which are organized into five fields at the base of each haltere. Despite the important role of these mechanosensory structures in flight, the central organization of the sensory afferents originating from the different field campaniforms has not been determined. We show here that in the blow fly, Calliphora vicina, sensory afferents from the campaniform fields project to the thorax in a region-specific manner. Afferents from different fields have different projection profiles and in addition, the projection pattern of afferents from different regions of the same field may show further variation. However, central target regions of these afferents are not exclusive to particular sensory fields because cells from different fields can possess similar projection profiles. Convergence of afferent projections is not limited to axons from the haltere fields, but is also observed between afferents originating from the haltere fields and those from serially homologous fields on the radial vein of the wing. Although we have not determined the specific cellular targets of the haltere sensory cells, the afferents of a dorsal field could make potential contact with at least one identified wing steering motoneuron that is known to be important in turning maneuvers. Our results, thus, provide the anatomical basis for studying how mechanosensory information encoded by the complex fields on the base of the haltere is mapped onto different functional regions within the CNS. © 1996 Wiley-Liss, Inc.     

Central afferent projections of proprioceptive sensory neurons in Drosophila revealed with the enhancer-trap technique

We have used a GAL4 enhancer-trap line coupled with an upstream activation sequence (UAS)-linked lacZ reporter construct to visualise and describe the central projections of proprioceptive sensory neurons of the thorax and abdomen in Drosophila. In the legs, lacZ expression is restricted to sensory neurons associated with hair plates, a subset of campaniform sensilla, and with the femoral chordotonal organ; whereas, in the wing, expression is seen only in subsets of campaniform sensilla. In the abdomen, expression is seen in Wheeler's organ and in a segmentally repeated array of internal sensory neurons that have not been previously described. The central projections from all of these neurons are described. The results confirm and expand upon our knowledge of the organisation of sensory neuropils in insects. The enhancer-trap technique provides a potentially powerful tool for describing the organisation of the central nervous system of Drosophila. © 1996 Wiley-Liss, Inc.     

Central projections of axons from taste hairs on the labellum and tarsi of the blowfly, Phormia regina Meigen.

Taste hairs are located on the labellum and tarsi of blowflies. These multimodal hairs consist of four functionally distinct chemoreceptors and a mechanoreceptor. By staining selected multimodal hairs, we sought to identify the central projection patterns of multiple and single axons from those hairs. On each side of the labellum there are 11 "largest" hairs (LH). The neurons associated with the anteriormost (LH-1), posteriormost (LH-11), and one lateral (LH-6) hair on the labellum were stained selectively with cobaltous sulfide. The overall projection pattern in the central nervous system (CNS) for axons from LH-1 and LH-11 is similar and differs markedly from axons from LH-6. At least three individual axon-projection patterns were determined for each labellar hair filled, indicating a partial functional organization for axons from multimodal hairs. One identified axon, the dorsalmost axon, has terminal arborizations that do not differ with the location of its associated hair. Another axon, thicker than the others, projects to a region that is distinct from the four thin axons. Within this region the arborizations of the thick axons occupy different areas depending on the location of their associated hair. Neurons from the largest hairs on the distalmost tarsomere (D5) of each leg were also stained and consisted of one thick and four thin axons. All axons except one thin axon from tarsal D5 hairs terminate in their respective leg neuromeres. The remaining thin axon projects to the suboesophageal ganglion ipsilateral to the hair filled and terminates in the same region as a branch of the labellar dorsalmost axon. These data suggest that axonal arbors from multimodal hairs have a limited functional and somatotopic organization in the blowfly CNS.     

Primary sensory ganglion cells projecting to the principal trigeminal nucleus in the mallard, Anas platyrhynchos

The trigeminal and glossopharyngeal ganglia of the adult mallard were studied following HRP injections into the principal trigeminal nucleus (PrV). The PrV consists of the principal trigeminal nucleus proper (prV) and the principal glossopharyngeal nucleus (prIX). After an injection into the prV, the labeled cells were found in the ipsilateral trigeminal ganglion. After an injection into the prIX, labeled cells were found in the ipsilateral distal glossopharyngeal ganglion, but not in the proximal ganglion of the IX and X cranial nerve (pGIX + X). In Nissl preparations, two types of ganglion cells in the trigeminal ganglion, pGIX + X, and distal ganglion of N IX could be distinguished: larger light cells and smaller dark cells. We could not determine whether the HRP-labeled cells belonged to both types or to one of them; but because all the labeled cells were over 20 microns, we concluded that the smallest cells (10-19 microns) in the trigeminal ganglion and distal ganglion of N IX did not project to the PrV. The labeling of the cells in the distal ganglion of N IX (average 34.5 microns) was uniformly moderate. In the trigeminal ganglion there were two types of labeled cells: heavily labeled cells (average 29.1 microns) and moderately labeled cells (average 35.1 l microns). These two types of labeling (moderate and heavy) may reflect two types of primary sensory neurons: cells with ascending, nonbifurcating axons, and cells with bifurcating axons. We speculate that the former are proprioceptive neurons and the latter tactile neurons. Labeled bifurcating axons in the sensory trigeminal complex gave off collaterals to all parts of the descending trigeminal nucleus except to the caudalmost laminated spinal part.     

Phylogenetic investigation of Dogiel's pericellular nests and Cajal's initial glomeruli in the dorsal root ganglion

Cajal's initial glomeruli (IG) and Dogiel's pericellular nests (PCNs) were first described from methylene blue preparations of healthy animal tissues around the beginning of the last century. Since that time, although many reports have been published concerning these structures, few have focused on their development and phylogeny in healthy animals. The aim of this study was to examine the phylogenetic development of the sensory neurons in Cajal's IG (also called axonal glomeruli) and Dogiel's PCNs in the dorsal root ganglion (DRG) of the healthy adult frog, chick, rat, and rabbit. The three-dimensional architecture of the neurons was observed in ganglia by scanning electron microscopy after removal of the connective tissue. The neurons in the DRG of fish are known to be bipolar, but DRG neurons in the species examined here were found to be pseudounipolar, with single stem processes. The proportion of neurons having IG or PCNs increased with increasing phylogenetic complexity in the species examined here. Cajal's initial glomeruli, the convolution of the stem process near the parent cell body: In frogs, the ganglia were small and the neuronal stem processes were very short and straight. In chicks, the stem processes were longer; sometimes very long, tortuous processes were observed. However, no neurons with typical IG were observed in either species. Typical IG were observed in rats and rabbits; their occurrence was much more frequent in rabbits. Pseudounipolarization, i.e., the transition from bipolar to pseudounipolar neurons, is thought to save space, limit the length of neuronal processes, and reduce conduction time. However, an explanation of the evolutionary advantage of the IG, which is formed by the excessive prolongation of the stem process, remains elusive. The cytological and electrophysiological importance of IG has been discussed. Dogiel's pericellular nests (PCNs), which resemble balls of yarn made of thin unmyelinated nerve fibers around DRG neurons, have been observed in the DRG of rats and rabbits, but not in frogs or chicks. This interesting structure shows not only ontogenetic development in healthy animals but also phylogenetic development among species. The nerve fibers in the PCNs were less than 1.2 mum in diameter and had some varicosities. An immunohistochemical study using anti-tyrosine hydroxylase (TH) antibody revealed that some PCNs contain TH-positive nerve fibers and varicosities. Such TH-positive PCNs disappear after sympathectomy. These results suggest that the PCNs are made up of autonomic nerve fibers.     

Area 3a in the cat. II. Projections to the motor cortex and their relations to other corticocortical connections.

It is well known that area 3a in the cat may monosynaptically influence the activity of neurons in the motor cortex. Much less information is available, however, on the anatomy of these connections. By using single or combined injections of different retrograde axonal tracers, we investigated the topography (horizontal and laminar) of area 3a neurons projecting to the motor cortex, and the anatomical relationships between these neurons and those projecting to other areas (2, 5, and SII) which, in turn, project to the motor cortex. Area 3a projects to all parts of area 4 gamma, to area 4 delta, and to the agranular area 6 in the lateral bank of the presylvian sulcus (area 6 alpha gamma), but not to other parts of areas 4 and 6. This projection exhibits a loose topographic organization along the mediolateral dimension of area 3a, and, in many cases, arises predominantly from the rostral half of this area. Although single small injections in the motor cortex produced two or more separate patches of retrograde labeling in 3a, after simultaneous injections of fluorochromes in two separate loci there often appeared in area 3a overlapping populations of neurons which were labeled retrogradely by each of the dyes, but with very few double-labeled neurons. In horseradish peroxidase (HRP) cases, 72% of area 3a neurons projecting to area 4 gamma were distributed in supragranular layers (mainly layer III), although the proportion of labeling in infragranular layers was larger when using fluorescent dyes. Double-labeled cells predominated in infragranular layers. These results have a bearing upon the functional roles that have been attributed to area 3a, as a cortical locus involved in muscle sensation, and a cortical relay to the motor cortex of rapid feedback information from muscle activity during movement.     

The ratio of dorsal root ganglion cells to dorsal root axons in sacral segments of the cat

Processes of dorsal root ganglion cells are depicted as being unbranched until they reach the spinal cord or periphery. This is an important concept because, for example, branching of these processes might be a basis for referred pain. Recently several studies indicate that axons of rat dorsal root ganglion cells branch in or near the ganglion. The present study extends this work by showing that there are approximately 1.4 dorsal root axons for each dorsal root ganglion cell in sacral segments of the adult cat, and these data are interpreted as indicating that many dorsal root axons in this animal also branch. Thus this study provides further evidence to indicate that a revision of our ideas about the organization of primary sensory cells is desirable. In addition, this study provides data to indicate that the numbers of both dorsal root ganglion cells and dorsal root axons differ on the right as compared to the left side of the same segment.     

Central anatomy of individual rapidly adapting low-threshold mechanoreceptors innervating the hairy skin of newborn mice: early maturation of hair follicle afferents.

Adult skin sensory neurons exhibit characteristic projection patterns in the dorsal horn of the spinal gray matter that are tightly correlated with modality. However, little is known about how these patterns come about during the ontogeny of the distinct subclasses of skin sensory neurons. To this end, we have developed an intact ex vivo somatosensory system preparation in neonatal mice, allowing single, physiologically identified cutaneous afferents to be iontophoretically injected with Neurobiotin for subsequent histological analyses. The present report, centered on rapidly adapting mechanoreceptors, represents the first study of the central projections of identified skin sensory neurons in neonatal animals. Cutaneous afferents exhibiting rapidly adapting responses to sustained natural stimuli were encountered as early as recordings were made. Well-stained representatives of coarse (tylotrich and guard) and fine-diameter (down) hair follicle afferents, along with a putative Pacinian corpuscle afferent, were recovered from 2-7-day-old neonates. All were characterized by narrow, uninflected somal action potentials and generally low mechanical thresholds, and many could be activated via deflection of recently erupted hairs. The central collaterals of hair follicle afferents formed recurrent, flame-shaped arbors that were essentially miniaturized replicas of their adult counterparts, with identical laminar terminations. The terminal arbors of down hair afferents, previously undescribed in rodents, were distinct and consistently occupied a more superficial position than tylotrich and guard hair afferents. Nevertheless, the former extended no higher than the middle of the incipient substantia gelatinosa, leaving a clear gap more dorsally. In all major respects, therefore, hair follicle afferents display the same laminar specificity in neonates as they do in adults. The widely held misperception that their collaterals extend exuberant projections into pain-specific regions of the dorsal horn during early postnatal life is shown to have multiple, deep-rooted underpinnings.     

Laminar Origin of Corticostriatal Projections to the Motor Putamen in the Macaque Brain

In the macaque brain, projections from distant, interconnected cortical areas converge in specific zones of the striatum. For example, specific zones of the motor putamen are targets of projections from frontal motor, inferior parietal, and ventrolateral prefrontal hand-related areas and thus are integral part of the so-called “lateral grasping network.” In the present study, we analyzed the laminar distribution of corticostriatal neurons projecting to different parts of the motor putamen. Retrograde neural tracers were injected in different parts of the putamen in 3 Macaca mulatta (one male) and the laminar distribution of the labeled corticostriatal neurons was analyzed quantitatively. In frontal motor areas and frontal operculum, where most labeled cells were located, almost everywhere the proportion of corticostriatal labeled neurons in layers III and/or VI was comparable or even stronger than in layer V. Furthermore, within these regions, the laminar distribution pattern of corticostriatal labeled neurons largely varied independently from their density and from the projecting area/sector, but likely according to the target striatal zone. Accordingly, the present data show that cortical areas may project in different ways to different striatal zones, which can be targets of specific combinations of signals originating from the various cortical layers of the areas of a given network. These observations extend current models of corticostriatal interactions, suggesting more complex modes of information processing in the basal ganglia for different motor and nonmotor functions and opening new questions on the architecture of the corticostriatal circuitry.SIGNIFICANCE STATEMENT Projections from the ipsilateral cerebral cortex are the major source of input to the striatum. Previous studies have provided evidence for distinct zones of the putamen specified by converging projections from specific sets of interconnected cortical areas. The present study shows that the distribution of corticostriatal neurons in the various layers of the primary motor and premotor areas varies depending on the target striatal zone. Accordingly, different striatal zones collect specific combinations of signals from the various cortical layers of their input areas, possibly differing in terms of coding, timing, and direction of information flow (e.g., feed-forward, or feed-back).     

Neural control of leg movements in a metamorphic insect: sensory and motor elements of the larval thoracic legs in Manduca sexta

During the metamorphosis of the hawkmoth Manduca sexta the larval thoracic legs degenerate to be replaced in the adult by legs of very different form and function. This change must be accompanied by a reorganization of the neural circuits controlling leg movements. As an initial step in the study of this reorganization we describe here the sensory and motor elements of this circuitry in the larval stage of life. Sensory neurons innervating mechanoreceptive hairs on the thoracic surface were stained individually with cobalt. Those innervating hairs on the general thoracic surface project topographically into two ventral regions of the segmental ganglia. Sensory neurons innervating leg sensilla also map topographically to the more ventral of these regions but in addition have arborizations in a midlateral region. The density of branching within this lateral "leg neuropil" is greatest for sensory neurons form sensilla on the more distal leg segments. Leg motor neurons were identified with intracellular recording and cobalt injection techniques. Those innervating muscles controlling distal leg segments have dense dendritic arbors in the lateral "leg neuropil," while motor neurons controlling more proximal segments and muscles of the ventral body wall have extensive arborizations in a dorsomedial region of the ganglion. In general, flexor motor neurons are excited by medial and inhibited by lateral leg sensilla, while the opposite is true of extensors. Distal segment motor neurons respond most strongly to sensory neurons from distal segments, thus suggesting some interaction within the lateral "leg neuropil." Thus, in the larval nervous system a highly ordered array of of sensory and motor elements underlies the specific behavioral responses of the legs to tactile stimulation.     

Projections of the second cervical dorsal root ganglion to the cochlear nucleus in rats

Physiological, anatomical, and clinical data have demonstrated interactions between somatosensory and auditory brainstem structures. Spinal nerve projections influence auditory responses, although the nature of the pathway(s) is not known. To address this issue, we injected biotinylated dextran amine into the cochlear nucleus or dorsal root ganglion (DRG) at the second cervical segment (C2). Cochlear nucleus injections retrogradely labeled small ganglion cells in C2 DRG. C2 DRG injections produced anterograde labeling in the external cuneate nucleus, cuneate nucleus, nucleus X, central cervical nucleus, dorsal horn of upper cervical spinal segments, and cochlear nucleus. The terminal field in the cochlear nucleus was concentrated in the subpeduncular corner and lamina of the granule cell domain, where endings of various size and shapes appeared. Examination under an electron microscope revealed that the C2 DRG terminals contained numerous round synaptic vesicles and formed asymmetric synapses, implying depolarizing influences on the target cell. Labeled endings synapsed with the stalk of the primary dendrite of unipolar brush cells, distal dendrites of presumptive granule cells, and endings containing pleomorphic synaptic vesicles. These primary somatosensory projections contribute to circuits that are hypothesized to mediate integrative functions of hearing.     

Golgi study of the primate substantia nigra. II. Spatial organization of dendritic arborizations in relation to the cytoarchitectonic boundaries and to the striatonigral bundle

The spatial organization of Golgi-stained dendritic arborizations of the substantia nigra was studied in three dimensions by using a video computer system. Dendritic orientation was analyzed in relation to the cytoarchitectonic boundaries and to the direction of the axons of the striato-pallidonigral bundle. All the brains, humans and macaques, were sectioned according to the same ventricular planes. The striatal bundle is made up of dense fascicles of very thin parallel axons. Sixty neurons located in the pars reticulata, lateralis, and compacta were reconstructed from serial sections. In the anterior pars reticulata and lateralis, the dendritic arborizations spread in all directions inside the striatal bundle. Below the pars compacta fringes, the dendrites of pars reticulata neurons extend ventrolaterally in the bundle. Because one nigral arborization can cover the whole thickness of the striatal bundle, we are led to believe that nigral neurons exert a role of convergence of the corticostriatal information similar to that of pallidal neurons (Percheron et al., '84a,b). The pars reticulata neurons appear to receive information mainly from the associative striatal territory. The pars lateralis neurons, conversely, appear to receive information from the sensorimotor territory. The anterior pars compacta neurons are organized in such a way that their ventral dendrites, located inside the pars reticulata, are ventrolaterally oriented, perpendicular to the striatal bundle. Their dorsal dendrites remaining in the pars compacta can receive other input. At more caudal levels, the posterior pars compacta neurons have dendrites radiating outside the striatal bundle.     

Focal myoclonus-dystonia of the leg secondary to a lesion of the posterolateral putamen: clinical and neurophysiological features.

We report on a patient with spontaneous and stimulus-sensitive myoclonic jerks and dystonia of the right leg that had been present since infancy. Magnetic resonance imaging showed a linear area of gliosis confined to the left posterolateral putamen. This is the first report of focal myoclonus-dystonia of the lower limb secondary to a putaminal lesion.     

Remodeling of a larval skeletal muscle motoneuron to drive the posterior cardiac pacemaker in the adult moth, Manduca sexta

During postembryonic development, a larval skeletal muscle motoneuron, MN-1 in abdominal segments 7 and 8, becomes respecified to innervate the terminal cardiac chamber of adult Manduca sexta. Neural tracing techniques and electrophysiology were used in this study to describe the anatomical and physiological remodeling of this identified motoneuron. During metamorphosis the MN-1 in segments 7 and 8 undergoes dendritic reorganization. Long new dendrites extend anteriorly in the terminal ganglion neuropil. Intracellular and extracellular recordings showed that broader action potentials, increased firing rate, and development of a bursting activity pattern accompany MN-1 respecification. Cardiac mechanograms showed that MN-1 activity bursts always correlate with the anterograde cardiac beat. Bilateral MNs-1 fire at similar times to activate and sustain the putative cardiac pacemaker activity of the terminal chamber synergistically. After remodeling, MN-1 output could be influenced rapidly by sensory inputs during evoked cardiac reversal. The effect is exerted by inhibition of MN-1 firing that, in turn, causes early blockade of the anterograde beat and reversal to the retrograde direction of beat.     

Projections from the nucleus accumbens to cholinergic neurons of the ventral pallidum: a correlated light and electron microscopic double-immunolabeling study in rat

A correlated light- and electron microscopic double-immunolabeling study combining choline acetyltransferase immunocytochemistry with anterograde tracing ofPhaseolus vulgaris leucoagglutinin (PHA-L) revealed that axons of the nucleus accumbens terminate on cholinergic neurons of the ventral pallidum. These findings are discussed with respect to the possibility that these cholinergic neurons may be part of parallel circuits, providing feedback to the same cortical and amygdaloid areas which innervate the nucleus accumbens.     

Developmentally Regulated Neurite Outgrowth Response from Dorsal Root Ganglion Neurons to Heparin-binding Growth-associated Molecule (HB-GAM) and the Expression of HB-GAM in the Targets of the Developing Dorsal Root Ganglion Neurites

Heparin-binding growth-associated molecule (HB-GAM) is a highly conserved cell surface- and extracellular matrix-associated protein that enhances neurite outgrowth in brain neurons in vitro. To study the possible response of peripheral neurons, we cultured chicken dorsal root ganglion neurons from different developmental stages from embryonic day 4.5 (E4.5; St 25) to E9 (St 35) on recombinant HB-GAM. We discovered that the neurite outgrowth response to HB-GAM is maximal at E5.5-6.5 (St 28-30). In order to correlate this in vitro phenomenon with in vivo phenomena, immunohistochemical staining and in situ hybridization were performed on cryosections. The protein expression of HB-GAM peaked at E6 (St 29) and was most extensive on the dorsal spinal cord and dorsal roots. Using Dil labelling, we confirmed that at the time when sensory afferents travel longitudinally in the bundle of His of the spinal cord, HB-GAM protein expression there is at its peak. Though HB-GAM is a secreted protein, at the RNA level the timing of HB-GAM appearance and existence in the spinal cord and sensory ganglia is in accordance with its protein expression. Our results demonstrate that peripheral neurons are responsive to substrate-bound HB-GAM in a developmentally regulated manner, and that the expression of both HB-GAM mRNA and protein in vivo is spatially and temporally matched to this in vitro phenomenon. HB-GAM is therefore a putative cue for the growth of sensory afferents to and within the dorsal spinal cord.     

Cholinergic systems in the rat brain: III. Projections from the pontomesencephalic tegmentum to the thalamus, tectum, basal ganglia, and basal forebrain

The ascending cholinergic projections of the pedunculopontine and dorsolateral tegmental nuclei, referred to collectively as the pontomesencephalotegmental (PMT) cholinergic complex, were investigated by use of fluorescent tracer histology in combination with choline-O-acetyltransferase (ChAT) immunohistochemistry and acetylcholinesterase (AChE) pharmacohistochemistry. Propidium iodide, true blue, or Evans blue was infused into the anterior, reticular, mediodorsal, central medial, and posterior nuclear areas of the thalamus; the habenula; lateral geniculate; superior colliculus; pretectal/parafascicular area; subthalamic nucleus; caudate-putamen complex; globus pallidus; entopeduncular nucleus; substantia nigra; medial septal nucleus/vertical limb of the diagonal band area; magnocellular preoptic/ventral pallidal area; and lateral hypothalamus. In some animals, separate injections of propidium iodide and true blue were made into two different regions in the same rat brain, usually a dorsal and a ventral target, in order to assess collateralization patterns. Retrogradely transported fluorescent labels and ChAT and/or AChE were analyzed microscopically on the same brain section. All of the above-delimited targets were found to receive cholinergic input from the PMT cholinergic complex, but some regions were preferentially innervated by either the pedunculopontine or dorsolateral tegmental nucleus. The former subdivision of the PMT cholinergic complex projected selectively to extrapyramidal structures and the superior colliculus, whereas the dorsolateral tegmental nucleus was observed to provide cholinergic input preferentially to anterior thalamic regions and rostral portions of the basal forebrain. The PMT cholinergic neurons showed a tendency to collateralize extensively.     

Cervical dorsal root ganglion cells with collaterals to both shoulder skin and the diaphragm. A fluorescent double labelling study in the rat. A model for referred pain?

Rat cervical spinal ganglion cell were retrogradely double-labelled with fluorescence dyes following injection of one dye into a cutaneous shoulder nerve and the other into the diaphragm. We suggest that the peripheral dichotomization of these ganglion cells could form the structural basis for the referred ‘phrenic’ pain in the shoulder region that can follow irritation of the diaphragm.