Mapping of neurons in the gravity receptor system of the octopus statocyst by iontophoretic cobalt staining

Summary The presence of uni-, bi- and multipolar neurons beneath the hair cell epithelium of the Octopus gravity receptor system has been demonstrated by iontophoretic cobalt staining. Counts give an average number of 1,940 neurons per macula. Whether the hair cells are primary of secondary sensory cells is discussed.This work was supported by grant Wo 160/3 of the Deutsche Forschungsgemeinschaft (DFG) to H.G.W. Thanks are due to the Director and staff of the Zoological Station in Naples for their hospitality and help     

The optic lobe projection pattern of polarization-sensitive photoreceptor cells in Drosophila melanogaster

Summary Histological staining of wild-type and sevenless transgenic Drosophila melanogaster bearing Rh3-lacZ fusion genes permits the selective visualization of polarization-sensitive R7 and R8 photoreceptor cells located along the dorsal anterior eye margin. Diffusion of -galactosidase throughout these cells reveals that they project long axons to the two most peripheral synaptic target rows of the dorsal posterior medulla, defining a specialized marginal zone of this optic lobe. Comparison of the staining patterns of marginal and nonmarginal Rh3-lacZ-expressing photoreceptor cells in the same histological preparations suggests that the marginal cells possess morphologically specialized axons and synaptic terminals. These findings are discussed with reference to the neuroanatomy of the corresponding dorsal marginal eye and optic lobe regions of the larger dipterans Musca and Calliphora, and in relation to the ability of Drosophila to orient to polarized light.     

Localization of corticotropin-releasing factor-immunoreactive nervous tissue and colocalization with neuropeptide Y-like substance in the optic lobe and peduncle complex of the octopus (<Emphasis Type="Italic">Octopus vulgaris</Emphasis>)

The distribution of corticotropin-releasing factor (CRF)-like immunoreactivity and its colocalization with neuropeptide Y (NPY)-like substances were investigated in the optic lobe and peduncle complex of the octopus (Octopus vulgaris) using immunohistochemical techniques. In the optic lobe cortex, CRF-immunoreactive (CRF-IR) and NPY-immunonegative varicose fibers were observed in the plexiform layer. In the medulla, CRF-IR somata were seen in the cell islands, and CRF-IR varicose fibers were observed in the neuropil. About half of the CRF-IR structures in the medulla showed NPY-like immunoreactivity. In the peduncle lobe, no CRF-IR somata but abundant CRF-IR varicose fibers were observed, and about half of them showed NPY-like immunoreactivity. In the olfactory lobe, CRF-IR somata and abundant CRF-IR varicose fibers were observed. Almost all the CRF-IR somata located in the posterior olfactory lobule showed NPY-like immunoreactivity, whereas those seen in the median olfactory lobule were immunonegative for NPY. About half of the CRF-IR fibers in the anterior lobule neuropil were immunopositive for NPY, but those in the median and posterior lobule neuropils were immunonegative for NPY. In the optic gland, almost all the CRF-IR varicose fibers were immunoreactive for NPY. Western blot analysis of the optic lobe and peduncle complex indicated that anti-CRF antiserum labeled approximate 16.4- and 14.6-kDa bands and that anti-NPY antiserum labeled an approximate 16.2-kDa band. CRF-IR and NPY-immunoreactive neurons in the optic lobe may participate in the modulation of visual information and those in the optic gland may be involved in the regulation of endocrine function.     

Two types of very long visual fibers found in the optic lobe of the flesh-fly,Boettcherisca peregrina

Summary Horseradish peroxidase (HRP) uptake (through a Corneal incision) in photoreceptor cells of the compound eye of Boettcherisca peregrina, resulted in the labeling of two types of very long visual fibers. One of them (the long fiber, If) penetrates through the lamina and medulla, and directly terminates within the lobula. The other (the bypass fiber, bpf) terminates in the medulla, like the axons of R7 and R8 photoreceptor cells, but the fibers run a considerably roundabout course. The photoreceptor cells with these very long fibers are region-dependent within the retina. Both the If and bpf are found almost exclusively in the male fly.     

The optic lobe of Drosophila melanogaster

Summary We present a quantitative evaluation of Golgiimpregnated columnar neurons in the optic lobe of wildtype Drosophila melanogaster. This analysis reveals the overall connectivity pattern between the 10 neuropil layers of the medulla and demonstrates the existence of at least three major visual pathways. Pathway 1 connects medulla layer M10 to the lobula plate. Input layers of this pathway are M1 and M5. Pathway 2 connects M9 to shallow layers of the lobula, which in turn are tightly linked to the lobula plate. This pathway gets major input via M2. Pathways 1 and 2 receive input from retinula cells R1-6, either via the lamina monopolar cell L1 (terminating in M1 and M5) or via L2 and T1 (terminating in M2). Neurons of these pathways typically have small dendritic fields. We discuss evidence that pathways 1 and 2 may play a major role in motion detection. Pathway 3 connects M8 to deep layers of the lobula. In M8 information converges that is derived either from M3 (pathway 3a) or from M4 and M6 (pathway 3b), layers that get their major input from L3 and R8 or L4 and R7, respectively. Some neurons of pathway 3 have large dendritic fields. We suggest that they may be involved in the computation of form and colour. Possible analogies to the organization of pathways in the visual system of vertebrates are discussed.During the final editing of this work our friend A.P.M. Dittrich was tragically killed in an accident. Without him this and the previous work would never have been completed     

The optic lobe of Drosophila melanogaster. I. A Golgi analysis of wild-type structure

Summary Golgi studies of the neurons in the optic lobes of Drosophila melanogaster reveal a large number of neuronal cell types. These can be classified as either columnar or tangential. Columnar elements establish the retinotopic maps of the lamina, medulla, and lobula-complex neuropiles. They are classified according to the position of their cell bodies, the number, width, and level of their arborizations, and their projection areas. Tangential elements are oriented perpendicularly to the columns. The arborizations of different tangential neurons are restricted to different layers of the optic neuropiles, within such layers their dendritic fields may span the entire retinotopic field or only part of it. The abundance of cell types inside each of the columnar units of the optic lobe is discussed with regard to its possible functional significance. By means of their stratified arborizations the columnar neurons form what appear to be multiple sets of retinotopically organized parallel information processing networks. It is suggested that these parallel networks filter different kinds of visual information and thus represent structurally separated functional subunits of the optic lobe. Such a parallel organization of visual functions increases the sites for function-specific gene actions and may explain the behavioral phenotypes of recently isolated structural mutants of the optic lobe.     

Notch signaling regulates neuroepithelial stem cell maintenance and neuroblast formation in Drosophila optic lobe development

Notch signaling mediates multiple developmental decisions in Drosophila. In this study, we have examined the role of Notch signaling in Drosophila larval optic lobe development. Loss of function in Notch or its ligand Delta leads to loss of the lamina and a smaller medulla. The neuroepithelial cells in the optic lobe in Notch or Delta mutant brains do not expand but instead differentiate prematurely into medulla neuroblasts, which lead to premature neurogenesis in the medulla. Clonal analyses of loss-of-function alleles for the pathway components, including N, Dl, Su(H), and E(spl)-C, indicate that the Delta/Notch/Su(H) pathway is required for both maintaining the neuroepithelial stem cells and inhibiting medulla neuroblast formation while E(spl)-C is only required for some aspects of the inhibition of medulla neuroblast formation. Conversely, Notch pathway overactivation promotes neuroepithelial cell expansion while suppressing medulla neuroblast formation and neurogenesis; numb loss of function mimics Notch overactivation, suggesting that Numb may inhibit Notch signaling activity in the optic lobe neuroepithelial cells. Thus, our results show that Notch signaling plays a dual role in optic lobe development, by maintaining the neuroepithelial stem cells and promoting their expansion while inhibiting their differentiation into medulla neuroblasts. These roles of Notch signaling are strikingly similar to those of the JAK/STAT pathway in optic lobe development, raising the possibility that these pathways may collaborate to control neuroepithelial stem cell maintenance and expansion, and their differentiation into the progenitor cells.     

Characterization of PDF-immunoreactive neurons in the optic lobe and cerebral lobe of the cricket, Gryllus bimaculatus

Pigment-dispersing factor (PDF) is a neuropeptide playing important roles in insect circadian systems. In this study, we morphologically and physiologically characterized PDF-immunoreactive neurons in the optic lobe and the brain of the cricket Gryllus bimaculatus. PDF-immunoreactivity was detected in cells located in the proximal medulla (PDFMe cells) and those in the dorsal and ventral regions of the outer chiasma (PDFLa cells). The PDFMe cells had varicose processes spread over the frontal surface of the medulla and the PDFLa cells had varicose mesh-like innervations in almost whole lamina, suggesting their modulatory role in the optic lobe. Some of PDFMe cells had a hairpin-shaped axonal process running toward the lamina then turning back to project into the brain where they terminated at various protocerebral areas. The PDFMe cells had a low frequency spontaneous spike activity that was higher during the night and was often slightly increased by light pulses. Six pairs of PDF-immunoreactive neurons were also found in the frontal ganglion. Competitive ELISA with anti-PDF antibodies revealed daily cycling of PDF both in the optic lobe and cerebral lobe with an increase during the night that persisted in constant darkness. The physiological role of PDF is discussed based on these results.     

Role of JAK/STAT signaling in neuroepithelial stem cell maintenance and proliferation in the Drosophila optic lobe

During Drosophila optic lobe development, proliferation and differentiation must be tightly modulated to reach its normal size for proper functioning. The JAK/STAT pathway plays pleiotropic roles in Drosophila development and in the larval brain, has been shown to inhibit medulla neuroblast formation. In this study, we find that JAK/STAT activity is required for the maintenance and proliferation of the neuroepithelial stem cells in the optic lobe. In loss-of-function JAK/STAT mutant brains, the neuroepithelial cells lose epithelial cell characters and differentiate prematurely while ectopic activation of this pathway is sufficient to induce neuroepithelial overgrowth in the optic lobe. We further show that Notch signaling acts downstream of JAK/STAT to control the maintenance and growth of the optic lobe neuroepithelium. Thus, in addition to its role in suppression of neuroblast formation, the JAK/STAT pathway is necessary and sufficient for optic lobe neuroepithelial growth.     

Secondary sensory cells in the gravity receptor system of the statocyst of Octopus vulgaris

Summary The presence of secondary sensory cells in the Octopus gravity receptor system has been demonstrated. In serial thin sections of the receptor cells (hair cells) no axons were found leaving the cells. Instead, synapses were observed with synaptic vesicles lying inside the receptor cells. Both data clearly indicate that the receptor hair cells represent secondary sensory cells. In addition, efferent contacts to the receptor cells could be confirmed.This work was supported in part by grant Wo 160/5 of the Deutsche Forschungsgemeinschaft to Prof. Dr. H.G. WolffThe experimental work was done in part at the Zoological Station in Naples and at the Sechenov Institute of Evolutionary Physiology and Biochemistry of the USSR Academy of Sciences (Laboratory of Prof. Dr. Ya.A. Vinnikov), Leningrad, USSR. The authors thank Prof. Vinnikov and Dr. Tsirulis for stimulating discussions     

Postembryonic development of Arg-Phe-amide-like and cholecystokinin-like immunoreactive neurons in the blowfly optic lobe

Summary The adult optic lobes of the blowfly Calliphora erythrocephala were found to be innervated by more than 2000 neurons immunoreactive to antisera raised against the neuropeptides FMRFamide, its fragment RFamide, and gastrin/cholecystokinin (CCK). All of the CCK-like immunoreactive (CCK-IR) neurons also reacted with antisera to RFamide, FMRFamide and pancreatic polypeptide. A few RFamide/FMRFamide-like immunoreactive (RF-IR) neurons did not react with CCK antisera; they reacted instead with antisera to Leu-enkephalin and Met-enkephalin-Arg⁶-Phe⁷. The RF-IR neurons are, thus, heterogeneous with respect to their contents of immunoreactive peptides. Two of the RF-IR neuron types innervating the adult optic lobes could be traced in their entirety only after following their postembryonic development, because of the complexity of the trajectories of the immunoreactive neuronal process in the adult insect. The majority of the cell bodies of the RF-IR and CCK-IR neurons lie within the optic lobes and are derived from imaginal neuroblasts of the inner and outer optic anlagen. Six of the peptidergic neurons are, however, metamorphosing larval neurons with their cell bodies in the central part of the protocerebrum. The full extent of immunoreactivitiy is not attained in some of the neurons until the late pupal or early adult stage. The larval optic center was also found to be innervated by neurons immuno-reactive with both RFamide and CCK antisera. The cell bodies of these RF-IR/CCK-IR neurons are located near the developing lamina (one on each side). In the 24 h pupa, the cell bodies of these neurons are still immunoreactive, but thereafter they cannot be immunolabeled apparently due to cell death or a change in transmitter phenotype.     

Distribution of GABA-like immunoreactive neurons in the optic lobes of Periplaneta americana

Summary Specific antisera against protein-conjugated -aminobutyric acid (GABA) were used in immunocytochemical staining procedures to study the distribution of the putative GABA-like immunoreactive neurons in the optic lobes of Periplaneta. GABA-like immunoreactive structures are evident in all three optic neuropil regions. Six different populations of GABAergic neurons, whose perikarya are grouped around the medulla, are found within the optic lobe. The number of these immunoreactive cells varies greatly and corresponds to the number of ommatidia of the eye. In the proximal part of the lamina, a coarse network of GABA-positive fibres is recognizable. These are the processes of large field tangential cells whose fibres pass through the distal surface of the medulla. A second fibre population of the lamina is made up of the processes of the centrifugal columnar neurons whose perikarya lie proximally to the medulla. The medulla contains 9 layers with GABAergic elements of variable immunoreactivity. Layers 1, 3, 5, 7 and 9 exhibit strong labelling, as a result of partial overlapping of the processes of centrifugal and centripetal columnar neurons, tangential fibres and/or lateral processes of perpendicular fibres and (possibly) processes of amacrines. A strong immunoreactivity is found in the proximal and distal layers of the lobula.     

Dissociation of photoreceptor cells from the pineal organ of the lamprey,Lampetra japonica

Summary Photoreceptor cells, nerve cells and supporting cells were dissociated from the pineal organ of the river lamprey, Lampetra japonica, by the use of 10 U/ml papain solution at 28° C for 20 min, followed by repeated trituration. With the aid of Nomarski interferencecontrast optics, photoreceptor cells, nerve cells and supporting cells were readily identified. Electron-microscopic examination revealed that isolated photoreceptor cells display an outer segment endowed with a few lamellar disks and connected to the inner segment (ellipsoid) via a connecting cilium. The structural features of the dissociated photoreceptor and supporting cells strongly resemble the morphology of the respective cellular elements in situ. We succeeded in culturing dissociated cells for time periods up to 48 h when the procedure described in detail was applied.     

Direct connections between the R7/8 and R1–6 photoreceptor subsystems in the dipteran visual system

Summary Musca and related flies have three main photoreceptor subsystems. The R1–6 group has short axons that terminate in the cartridges of the first optic neuropile, the lamina. The cartridges are bypassed by the longer axons of R7 and R8, which run together to terminate at different levels in the underlying medulla neuropile. The present account describes a shallow, previously unidentified zone in the lamina within which R7/8 make glancing contact with R1–6. At the distal border of the cartridge over no more than 3–4 m depth, the tangentially directed short axon of R6 squeezes between the pair from R7 and R8, forming quite large areas of mutual contact (approximately 7 m²). Less frequently, R1 is contacted. At least some of these sites contain smaller membrane specialisations indistinguishable from the more numerous gap junctions found more proximally that interconnect the terminals of R1–6. The R7/8 junctions with R6 are of comparable size (0.15 m²) and likewise possess symmetrical membrane densities. They provide proposed pathways for direct electrical interaction to account for observed electrical input from R7/8 to the R1–6 subsystem. In two cases R7/8 was possibly postsynaptic to R1–6 at a multiple-contact synapse, but even if functional, these sites were so rare that they are unlikely to have much operational significance.     

Structural interactions of actin filaments and endoplasmic reticulum in honeybee photoreceptor cells

Summary Fluorescent phallotoxins and heavy meromyosin were used to reveal the organization of the actin cytoskeleton in honeybee photoreceptor cells, and the relationship of actin filaments to the submicrovillar, palisade-like cisternae of the endoplasmic reticulum (ER). Bundles of unipolar actin filaments (pointed end towards the cell center) protrude from the microvillar bases and extend through cytoplasmic bridges that traverse the submicrovillar ER. Within the cytoplasmic bridges, the filaments are regularly spaced and tightly apposed to the ER membrane. In addition, actin filaments are deployed close to the microvillar bases to form a loose web. Actin filaments are scarce in cell areas remote from the rhabdom; these areas contain microtubule-associated ER domains. The results suggest that the actin system of the submicrovillar cytoplasm shapes the submicrovillar ER cisternae, and that the distinct ER domains interact with different cytoskeletal elements.     

Types and arrangements of neurons in the crayfish optic lamina

Cell and Tissue Research - The neural arrangements in the optic lamina of the crayfish Pacifastacus leniusculus Dana have been studied by light microscopy by means of silver impregnation...     

Fine structural and volumetric changes of lamprey photoreceptor cells during light and dark periods

Summary Short photoreceptor cells of the lamprey retina exhibited a 30% increase in the width of the myoid process and a 20% increase in that of the axonal process during a 12-h light period, compared to the measurements obtained during a 12-h dark period. An increase in the amount of cytoplasm, dilation of ER cisterns, and swelling of the nucleus appeared to cause the enlargement of the myoid parts. Accumulation of synaptic vesicles occurred concurrently with a thickening of the axonal process. These morphological changes presumably represent a phase of the diurnal cycle and current synaptic activity of the short cell. By contrast, the long photorecpetor cell showed neither measurable changes nor any indication of retinomotor movement.     

Ultrastructure of the compound eye and first optic neuropile of the photoreceptor mutant ora JK84 of Drosophila

Summary The developmental mutant of Drosophila (ora ^JK84) is characterized by nonfunctional photoreceptor cells (R1–6), while the R7/R8 cells are normal. A fundamental question is: Does the near absence of photosensitive membranes inhibit development of the Rl-6 axons and their synapses at the other end of the cell? The retina and first optic neuropile (lamina ganglionaris) were examined with freeze-fracture technique and high voltage electron microscopy. R1–6 have reduced rhabdomere caps; rhabdomeric microvilli have about 50% of the normal diameter and 20% of the normal length. Affected cells exhibit prominent vacuoles which appear to communicate with some highly convoluted microvillar membranes. Almost no P-face particles (putative rhodopsin molecules) are present in the R1–6 rhabdomeres, and particle densities are lower in R7 than previously reported. Near the rhabdomere caps, microvilli of R1–6 are fairly normal, but at more proximal levels they are greatly diminished in length and changed in orientation, while at still more proximal levels they are lost. R1–6, R7, and R8 axons from each ommatidium are bundled into normal pseudocartridges beneath the basement membrane. No abnormalities are found in the lamina ganglionaris, and all synaptic associations as well as the presumed virgin synapses (of R1–6) appear normal. No glial anomalies are present, and R7/R8 axons project through the lamina in the usual fashion. These fine structural findings are correlated with known electrophysiological, biochemical, and behavioral correlates of both sets of photoreceptors (R1–6, and R7/R8).This study was supported substantially by the UW-HVEM Laboratory, in addition to a Faculty Development Award, a UMC Biomedical Research Support Grant N.I.H. RR07053 to W.S.S., and a Hatch Grant, Project 2100 to S.D.C. Freeze fracture was done at the Wisconsin Regional Primate Research Center, N.I.H. Grant RR00167. We thank Professor Hans Ris, Dr. J. Pawley, Dr. D. Neuberger, and Ms. M. Bushlow, HVEM Laboratory, Dept. of Zoology, UW. We also thank Mrs. K. Srivastava, Mr. M.B. Garment, Mr. G. Gaard, and Mr. D. Liu for technical assistance.     

Immunolocalization of Na,K-ATPase in blowfly photoreceptor cells

The Na,K-ATPase (sodium pump) plays a central role in the physiology of arthropod photoreceptors as it re-establishes gradients for Na⁺ and K⁺ after light stimulation. We have mapped the distribution of the Na,K-ATPase in the photoreceptors of the blowfly (Calliphora erythrocephala) by immunofluorescent and immunogold cytochemistry, and demonstrate that the distribution pattern is more complex than previously presumed. High levels of sodium pumps have been detected consistently in all photoreceptors R1-8 on the nonreceptive surface, but no sodium pumps are found on the microvillar rhabdomere. Within the nonreceptive surface of the cells R1-6, however, the sodium pumps are confined to sites juxtaposed to neighboring photoreceptor or glial cells; no sodium pumps have been detected on the parts of the nonreceptive surface exposed to the intra-ommatidial space. In R7 and R8, the sodium pumps are found over the entire nonreceptive surface. The cytoskeletal protein spectrin colocalizes with the sodium pumps suggesting that linkage of the pump molecules to the spectrin-based submembrane cytoskeleton contributes to the maintenance of the complex pattern of pump distribution.     

Cone photoreceptor shedding in the tree shrew (Tupaia belangerii)

Summary Tree shrews were sacrificed at various times during a 12 h light-12 h dark cycle and the retinal pigment epithelium (RPE) was examined for phagosomes. Analysis of photoreceptor densities showed that the tree-shrew retina consists of approximately 96% cone photoreceptors. Therefore, phagosomes in the RPE were assumed to be mostly those of cones. A peak in the number of RPE phagosomes was found about one hour after the onset of light. The number of phagosomes/mm RPE during the light cycle varied from 17.02 at the peak to 2.49 ten hours after light onset. During the dark cycle, values ranged from 0.10 to 0.61 phagosomes/mm RPE. Size profiles of phagosomes showed that large phagosomes peak in number 1/2 h after light onset, while smaller sizes peak at about 1 h after light onset. This may indicate that maximal shedding and phagocytotic activity occurs sometime before the peak in the total number of phagosomes is reached. Statistical corrections for phagosome size, section thickness and phagosomal degradation time were applied to the data in order to assess outer segment renewal time for tree shrew cones.