Sensory projections from normal and homoeotically transformed antennae in Drosophila

The sensory projection of homoeotic tarsal neurons in the antennal mutant spineless-aristapedia (ss^a) is compared with the projections of wild-type antennae and tarsi. The projection pattern was identified by diffusion of cobalt into the cut peripheral nerves followed by Timm's silver intensification. No sensory fibers of the homoeotic tarsus extend into the thoracic leg centers; instead they project into normal antennal centers of the brain. In the posterior antennal center and the posterior part of the suboesophageal ganglion (SOG) they show precisely the same pattern as do those from the wild-type antenna. In other regions this is not the case: in the antennal glomeruli homoeotic terminals are randomly distributed, and in the anterior SOG fibers form a tract which is not present in antennal cobalt fills. We have not found any correspondence between thoracic and homoeotic tarsal projections. The projection of homoeotic tarsi in mosaic flies exhibiting an ss^a antenna and a wild-type brain is similar to the “normal” homoeotic pattern. This suggests that the central nervous system (cns) is not transformed by the ss^a gene. The behavior of normal and ectopic sensory fibers in the cns is explained in terms of both intrinsic properties of the sensory axons and extrinsic factors in the surrounding nervous tissue.     

Surgical generation of supernumerary appendages for studying neuronal specificity in Drosophila melanogaster

The ability of sensory neurons to establish specific synaptic contacts in the central nervous system (CNS) can be studied by changing the spatial relationship between the periphery and the CNS. In contrast to the genetic displacement of appendages by homoeotic mutations, the surgical approach used in this study allows one to place homologous as well as heterologous appendages to the same site on the body surface. Using an improved technique of “surface transplantation,” we generated supernumerary appendages of any desired type in a particular abdominal position. The sensory axons originating from these grafts enter the CNS through the main abdominal nerve and arborize in the fused abdominal ganglia; many fibers extend also into thoracic centers. In the abdominal ganglia, terminals from dorsal transplants (wings and halteres) stay on the ipsilateral side, whereas terminals from ventral transplants (legs and antennae) distribute ipsi- and contralaterally. The same preference holds true for dorsal and ventral abdominal bristles, respectively, whose projection patterns served as a reference. In thoracic ganglia, axons from dorsal and ventral grafts yield completely different terminal patterns. Dorsal grafts project into the ipsilateral wing center, even in the mutant wingless, in which normal wing afferents are suppressed. In contrast, fibers from ventral grafts often extend along the thoracic midline. These data indicate that sensory axons of homologous appendages on the one hand, and their central targets on the other, share serially repeated surface markers. This may enable sensory fibers to recognize centers of homologous appendages.     

Directional sensitivity of an identified wind-sensitive interneuron during the postembryonic development of the locust

Simultaneous extracellular recordings from both locust abdominal connectives show a differential activation of both bilateral homologues of an identified long projection interneuron (A4I1) in response to wind stimuli from different directions. Despite the previously shown extensive structural dynamics of sensory afferents and synaptic rearrangement of the direct afferent-to-interneuron connections during postembryonic development, a directional sensitivity is already present in first instar nymphs. Only quantitative changes in the strength of the directional response can be detected. Intracellular stainings of the A4I1 interneuron in first instar nymphs and adults show that general morphological features do not change during postembryonic development, in contrast to the presynaptic sensory afferents. This also holds for general morphological features of pleuroaxillary flight motoneurons. The output connections of A4I1 to these motoneurons and an unidentified intersegmental interneuron are already present in flightless nymphs.     

Pattern formation and determination in the antenna of the homoeotic mutant Antennapedia of Drosophila melanogaster

The development of the antenna in the antennal-leg homoeotic mutant Antennapedia (Antp^R) was investigated using somatic crossing-over to mark clones of cells in Antp^R antennal appendages. Antp^R antennae ranged from a nearly normal antenna to a nearly normal leg. The arrangement of clones of marked bristles and cuticle in the more antennalike antennae was similar to the wild type antenna, and that of the leglike antennae was similar to the wild-type leg. The contiguity of clones argued against extensive individual cell migration. The regions occupied by homoeotic leg varied considerably between different Antp^R antennae. Observation of Antp^R antennae in these phenotypic mosaics showed that specific leg parts replaced specific antennal parts. Even small groups of leg sensilla appeared only at precise locations in the antenna. These results suggest that homoeotic leg cells and antennal cells can both respond to the same positional information or prepattern. An analysis of clone size provided estimates for cell number in the Antp^R antenna. It was found that cell numbers in the wild-type and Antp^R antennae are about the same until the third instar, when the Antp^R cells start dividing more rapidly than wild type. Previous work had shown that clonal inheritance of a commitment for homoeotic leg also did not occur prior to the early third instar. It is suggested that determination for homoeotic leg occurs in the early third instar, and that thereafter this commitment is inherited by the progeny of the determined cells. The increase in growth rate is probably due to a faster growth rate in cells with a leg commitment than in cells with an antennal commitment. The results suggest that, once initiated, determination may be of two types—a clonally inherited determination (for example, to be homoeotic leg) and an environmental determination (for example, to be a specific part of a homoeotic leg). Clonal inheritance of determination in normal embryonic development and in sex determination in intersexes is discussed.     

The Inductive Role of Bacterial Symbionts in the Morphogenesis of a Squid Light Organ

SYNOPSIS. The association of the sepiolid squid Euprymna scolopeswith its marine luminous bacterial symbiont Vibrio fischeriis an emerging model system to study the initiation and developmentof bacterial symbioses in higher animals, in particular theinfluence of bacteria on the ontogenic development of symbiotic-specifichost tissues. Experiments comparing the development of juvenilesquid infected with symbiotic V. fischeri with that of uninfectedjuveniles suggest postembryonic development of the light organrequires cell-cell interactions with the bacterial symbionts.The presence of symbiotic bacteria induces specific morphologicalchanges by affecting such fundamental processes as cell deathand cell differentiation. The surface of the juvenile organis largely composed of ciliated cells that appear to facilitateinfection of the light organ. These cells begin to undergo celldeath within hours of infection with symbiotic V. fischeri.Within three days the epithelial cells that form the bacteriacontainingcrypts of the light organ increase in size; these cells do notappear mitotically active, and may represent a terminally differentiatedstate. The light organs of uninfected juvenile E. scolopes,however, do not exhibit any of these early postembryonic developmentalevents but remain in a state of arrested morphogenesis.     

Central projections of a homoeotic regenerate, antennapedia, in a stick insect, Carausius morosus (Phasmida)

Homoeotic appendages provide a system for the analysis of neural path-finding in which the appendage is mismatched with its segmented ganglion. Central projections of sensory neurons from homoeotic antennapedia regenerates induced by antennal amputation in the stick insect, Carausius morosus, are described. The majority of afferent axons project to the olfactory lobe as in the normal antennal nerve, but they do not give rise to compact glomeruli. Nor does the form of the projection resemble that of leg sensory nerves in thoracic ganglia. The projection of antennapedia regenerate neurons in Carausius resembles the antennapedia mutant of Drosophila except that some primary afferents bypass the olfactory lobe and take several courses through the brain, sometimes reaching distant contralateral areas. It appears that these wandering fibers, having bypassed the olfactory lobe, tend to follow established tracts and to arborize or to deviate at circumscribed synaptic areas. The behavioral evidence for sensory input from antennapedia regenerates is equivocal.     

The effects of transplantation and regeneration of sensory neurons on a somatotopic map in the cricket central nervous system

The restoration of the cercal afferent projection of crickets was examined after severing the cercal nerve or amputating the cercus and reimplanting it. After either maneuver the sensory neurons regenerated arborizations in the central nervous system (CNS) within about 1 month. In order to assess the role of the pathway taken to the CNS in controlling the growth of the terminal arborization, we transplantated left cerci to the right side of the host. The operation mismatched the mediolateral axes of host and graft tissues. In one-third of the neurons examined, the axon trajectories of the regenerated neurons were altered. The terminal arborizations in these cases were unusual; for example, one neuron arborized in an abnormal area as well as in its normal area. In rare instances this neuron arborized only in incorrect areas of the CNS. Thus, it appears that axon pathway can have an effect on the central structure of sensory neurons. However, in most cases after the surgery, the neurons were able to reach their proper target areas even by circuitous routes. The proximodistal coordinate of the map is isomorphic with sensory neuron age, because the most distal receptors are produced early in postembryonic development and new ones are added proximally at each molt. We tested the possibility that the order of differentiation was critical for generating the afferent projection with two experiments. First, the distal cercus including the distal members of the clavate array was amputated. The specimen regenerated an entire distal cercus including distal clavate receptors. When newly generated, distal neurons were stained, the terminal arbors were identical to the amputated neurons they replaced. In this case, both age and order of arrival were reversed from normal yet the topographic projection pattern was not altered. Second, we transplanted young cerci onto older specimens and then examined the regenerated arbors of the transplanted sensory neuron. The immature neuron arborized in the adult nervous system exactly as the mature homolog. Thus the age of a sensory neuron did not appear to be a controlling variable in the elaboration of a terminal arborization. The significance of these results is discussed in the context of two models for development of orderly neuronal connections.     

Connectivity of identified central synapses in the cricket is normal following regeneration and blockade of presynaptic activity

Cercal sensory neurons in the cricket innervate interneurons in the central nervous system (CNS) and provide a model system for studying the formation of central synapses. When axons of the sensory neurons were transected during larval development, the cell bodies and the soma-bearing portion of axons, which are located within the cercus, survived but lost their excitability for 9-10 days. During this period, the sensory neurons grew new axons and reinnervated the terminal abdominal ganglion. Physiological recordings showed that sensory neurons of known identity reestablished monosynaptic contacts with their normal postsynaptic interneuron. Moreover, each synapse exhibited a characteristic strength indistinguishable from the intact synapse in an unoperated cricket. Since this selective connectivity was apparent immediately after the excitability of the axotomized sensory neurons was restored, action potentials in the sensory neurons appear to be unnecessary for normal synaptic regeneration to occur. Consistent with this, the reinnervation process was unaffected even when action potentials in the sensory neurons were blocked by tetrodotoxin (TTX) immediately following axotomy until just before testing. During the normal course of development, the characteristic strength of individual synapses changes systematically, resulting in the developmental rearrangement of these synapses (Chiba et al., 1988). This synaptic rearrangement was also unaffected when action potentials in the sensory neurons were blocked by TTX for the last 30% of larval development. Therefore, in the cricket cercal sensory system, both regeneration of the central synapses following axotomy of the presynaptic sensory neurons and the normal rearrangement of connectivity during larval development appear not to require axonal action potentials.     

Newborn cells in the adult crayfish brain differentiate into distinct neuronal types

Mitotically active regions persist in the brains of decapod crustaceans throughout their lifetimes, as they do in many vertebrates. The most well-studied of these regions in decapods occurs within a soma cluster, known as cluster 10, located in the deutocerebrum. Cluster 10 in crayfish and lobsters is composed of the somata of two anatomically and functionally distinct classes of projection neurons: olfactory lobe (OL) projection neurons and accessory lobe (AL) projection neurons. While adult-generated cells in cluster 10 survive for at least a year, their final phenotypes remain unknown. To address this question, we combined BrdU labeling of proliferating cells with specific neuronal and glial markers and tracers to examine the differentiation of newborn cells in cluster 10 of the crayfish, Cherax destructor. Our results show that large numbers of adult-generated cells in cluster 10 differentiate into neurons expressing the neuropeptide crustacean-SIFamide. No evidence was obtained suggesting that cells differentiate into glia. The functional phenotypes of newborn neurons in cluster 10 were examined by combining BrdU immunocytochemistry with the application of dextran dyes to different brain neuropils. These studies showed that while the majority of cells born during the early postembryonic development of C. destructor differentiate in AL projection neurons, neurogenesis in adult crayfish is characterized by the addition of both OL and AL projection neurons. In addition to our examination of neurogenesis in the olfactory pathway, we provide the first evidence that adult neurogenesis is also a characteristic feature of the optic neuropils of decapod crustaceans.     

Experimental analysis of sensory nerve pathways inDrosophila

Summary The pathway of adult sensory nerves has been analysed in three experimental situations: (i) in flies with grossly abnormal thoracic morphology resulting from X-irradiation early during development, (ii) in flies which had been subjected to surgical operations late in the larval period, (iii) in homoeotic mutants. The results provide experimental support for a simple mechanism in which developing adult axons join the nearest larval nerve and are guided by it up to the central nervous system. In particular, experimental interference with normal development can result in nerves from different segments, or from dorsal and ventral appendages, joining each other and entering the central nervous system together.     

Genetically displaced sensory neurons in the head of Drosophila project via different pathways into the same specific brain regions

The sensory projection pattern of homoeotic antennal or leg tissue in the proboscis of the mutant proboscipedia in Drosophila melanogaster was studied using orthograde diffusion of cobalt chloride. Both kinds of ectopic nerve fibers terminate in the normal center of the proboscis and in the antennal glomerulus of the brain. These centers are attained by axons which may pass via four different peripheral nerves. In contrast, wild-type proboscis fibers use only one pathway. Depending on the site of entrance into the brain, homoeotic axons may project into the proboscis center first, and then into the antennal glomerulus, or vice versa. Consequently, the tract between these two brain regions may be followed in opposite directions. Ectopic leg axons extending into normal leg centers in the thoracic ganglion have not been found. Projection patterns in proboscipedia are discussed together with those of the leg-like antenna in spineless-aristapedia (R. F. Stocker and P. A. Lawrene, 1981, Dev. Biol.82, 224–237). The fact that displaced antennal and leg neurons project specifically into normal proboscis and antennal centers may reflect the serial homology of antennal, leg, and proboscis neurons, and similar homology of the corresponding centers.     

Genetic Analysis of the Antennapedia Gene Complex (Ant-C) and Adjacent Chromosomal Regions of DROSOPHILA MELANOGASTER. II. Polytene Chromosome Segments 84A-84B1,2

The existence of a gene complex in the proximal right arm of chromosome 3 of Drosophila melanogaster involved in the development of the head and thorax was originally suggested by the phenotypes of several dominant homoeotic mutations and their revertants. A screen for mutations utilizing Df(3R) Antp^Ns+R17 (proximally broken in salivary region 84B1,2) yielded, among 102 recovered mutations, 17 localized by deficiency mapping to the putative homoeotic cluster. These fell into four complementation groups, two of which were characterized by homoeotic phenotypes. To explore the limits of the Antennapedia gene complex (ANT-C) more proximally, a second screen has been undertaken utilizing Df(3R)Scr, a deficiency of 84A1–B1,2.—Of 2832 chromosomes screened, 21 bearing alterations localized to polytene interval 84A–84B1,2 have been recovered. Sixteen are recessive lethals, and five showing reduced viability display a visible phenotype in surviving individuals. Complementation and phenotypic analyses revealed four complementation groups proximal to those identified in the previous screen, including two new alleles of the recessive homoeotic mutation, proboscipedia (pb). Ten of the new mutations correspond to complementation groups defined previously in the Df(3R)Antp^Ns+R17 screen four to the EbR11 group, two to the Scr group and four to the Antp group.—On the basis of the phenotypes of the 39 mutations localized to this region, plus their interactions with extant homoeotic mutations, we postulate that there are at least five functional sites comprising the ANT-C. Three have been demonstrated to be homoeotic in nature. The specific homoeotic transformations thus far observed suggest that these loci are critical for normal development of adult labial, maxillary and thoracic structures.     

Changes in the Inflammatory Response to Injury and Its Resolution during the Loss of Regenerative Capacity in Developing Xenopus Limbs

Tissue and organ regeneration, unlike development, involves an injury that in postembryonic animals triggers inflammation followed by resolution. How inflammation affects epimorphic regeneration is largely uninvestigated. Here we examine inflammation and its resolution in Xenopus laevis hindlimb regeneration, which declines during larval development. During the first 5 days postamputation, both regeneration-competent stage 53 and regeneration-deficient stage 57 hindlimbs showed very rapid accumulation of leukocytes and cells expressing interleukin-1β and matrix metalloproteinase 9. Expression of genes for factors mediating inflammatory resolution appeared more persistent at stages 55 and 57 than at stage 53, suggesting changes in this process during development. FoxP3, a marker for regulatory T cells, was upregulated by amputation in limbs at all three stages but only persisted at stage 57, when it was also detected before amputation. Expression of genes for cellular reprogramming, such as SALL4, was upregulated in limbs at all 3 stages, but markers of limb patterning, such as Shh, were expressed later and less actively after amputation in regeneration-deficient limbs. Topical application of specific proinflammatory agents to freshly amputated limbs increased interleukin-1β expression locally. With aqueous solutions of the proinflammatory metal beryllium sulfate, this effect persisted through 7 days postamputation and was accompanied by inhibition of regeneration. In BeSO₄-treated limbs expression of markers for both inflammation and resolution, including FoxP3, was prolonged, while genes for cellular reprogramming were relatively unaffected and those for limb patterning failed to be expressed normally. These data imply that in Xenopus hindlimbs postamputation inflammation and its resolution change during development, with little effect on cellular dedifferentiation or reprogramming, but potentially interfering with the expression of genes required for blastema patterning. The results suggest that developmental changes in the larval anuran immune system may be involved in the ontogenetic loss of epimorphic regeneration in this system.     

Developmental Aspects of Soybean (Glycine max) Somatic Embryogenesis

A detailed study of the developmental aspects for soybean somaticembryogenesis was undertaken with emphasis on biochemical andhistological markers. The various stages of somatic embryo developmentin callus cultures have been identified and characterized. Germinatingembryos could be converted to fertile plants at a high frequency(90%). Dicamba (3, 6-dichloro-o-anisic acid) was found to bethe auxin of choice for the clear distinction of the variousdevelopmental phases of soybean somatic embryos. Differencesin their protein patterns were determined using polyacrylamidegel electrophoresis. This analysis revealed distinguishabledifferences in protein profiles amongst the various developmentalstages, especially in heart stage embryos. Histological studieson somatic embryos revealed specific tissue types which closelyresemble those reported for zygotic embryos. Further evidenceis provided that there is a close similarity in tissue differentiation,between somatic and zygotic embryogenesis although there aresome unique features in the development of somatic embryos. Glycine max, callus cultures, developmental stage, liquid cultures, neomorphs, plant regeneration, stage specific proteins, histology     

The complete postembryonic development of Tropodiaptomus informis Kiefer, 1936 (Copepoda:Calanoida) reared in the laboratory

The complete postembryonic development of Tropodiaptomus informiscomprises six naupliar and six copepodid stages of which thelast is the adult. Of all the characteristics, it is the shapeand number of setae on the terminal segments of the antennulesin the late naupliar stages, and the shape and setae of exo-and endopods of the fifth legs from copepodids III to V whichare most useful in constructing relationships among eight diaptomidsreared.     

The Cardiac Mutant: An Overview

SYNOPSIS.The cardiac lethal mutation was discovered in a strainof axolotls (Ambysloma mexkanum) imported from Mexico. Embryoshomozygous recessive for the c gene develop in a normal fashionuntil the heartbeat stage of development. At this time, heartaction is observed in unaffected siblings, but in cardiac lethalembryos heart action is not apparent. Affected animals hatchand swim, but do not feed. Within a few days, ascites developsand the animals die within a short time afterward. The c geneis inherited as a simple recessive. The direct effect of gene c is unknown. Data has been presentedwhich suggests that gene c alters the anterior endoderm whichmay be a specific heart inductor in this species. Alternatively,the failure of heart action may result from an ionic imbalancein the immediate heart environment. This paper reviews the relevantexperiments and presents new data supporting each of these suggestionsfor the mechanism of action of the cardiac lethal gene.     

Genetic analysis ofloboid-ophthalmoptera,a homoeotic strain inDrosophila melanogaster

A homoeotic strain in which wing-like outgrowths are produced from the eye region was investigated. The actual wing nature of the malformation was confirmed. Genetically, this homoeotic character depends on the presence of an eye-reducing factor,loboid (ld; 3–102), and of modifier genes. One important sex-linked modifier, probably located at 1–5±, is thought to be the main factor underlying the homoeotic effect. It is designated asopht (ophthalmoptera), and the mutant strain asld-opht Kobel (1968), assuming a specific allele ofld with homoeotic effect, originally described this strain under the nameld ^oph. It is shown that inld-opht, theld factor can be replaced byDfd ^r-L (3–47.5) without irrevokably losing the homoeotic effect.Penetrance and expressivity ofopht are very variable and subject to genetic and environmental changes, and they readily respond to selection. Such properties are common to all homoeotic mutants. The phenomenon of homoeosis is interpreted in terms of allotypic differentiations resulting from a switching of development into other epigenetic pathways. This switching is perhaps due to an altered rate of proliferation.Aided by a grant from the Netherlands Organization for the Advancement of Pure Research (Z.W.O.).     

Characterisation of a new tumorous-head mutant of Drosophila melanogaster

Summary A new homoeotic mutant, I127, showing abnormal growths in the head region including homoeotic transformation of eye to genitalia and antenna to leg, was isolated in a screen designed to find new alleles of the tumorous head (tuh-3), mutation. Similarities in the phenotype and genetics of the mutant, and complementation studies with tuh-1; tuh-3, suggest that I127 is indeed an allele of tuh-3. In combination with the first chromosome modifier tuh-1, the mutant is temperature-sensitive during the third larval instar, giving an increased penetrance of the tumorous head phenotype when reared at 25° C as opposed to 18° C. The isolation of further alleles at the tumorous-head locus are essential. The types of morphological defects which can result from mutations at this locus would enable us to establish if this is a complex locus, and if null mutations are lethal during development. The interactions of the tumorous-head gene with first chromosome modifiers and other homoeotic mutations will only be understood if we able to induce a number of mutations at this locus, and as a consequence begin to elucidate the role of the wild-type gene product in normal development.