Morphological studies of possible neuroendocrine structures in Helisoma tenue (Gastropoda: Pulmonata)

Summary Studies of possible neuroendocrine structures in the pulmonate gastropod Helisoma tenue show that cerebral fuchsinophilic neurons with electron-dense granules (mean diameter 1,500 Å) apparently release their secretory content in an intercerebral commissure neurohemal area near the mediodorsal bodies (MDB) or in the median labial nerve neurohemal area.The MDB cells have axon-like processes which branch and end at the cerebral surface, separated by a thin capsule from the cerebral fuchsinophilic cells. The presence of granules (mean diameter 800 Å) in the terminals of the MDB cells suggests cell body origin, distal transport and release of the granular secretory material. The secretory product may have an influence on cerebral fuchsinophilic neurons.Electron microscopy reveals the presence of granules of different sizes and densities in expanded neurites at the periphery of the intestinal nerve of the visceral ganglion which may indicate the presence of a neurohemal area. However, the granules in the intestinal nerve neurites and in the visceral ganglion fuchsinophilic cells are similar to granules found in the heart which also suggests that the granules may carry a neurotransmitter.This study was supported by NTH traineship USPHS T1 CA 5045 and NSF grant GB-6424X. The author would like to thank Professor Howard A. Bern for guidance and support during the course of this work, Mr. John Underhill and Mr. Albert Blaker for photographic assistance, and Mrs. Emily Reid for preparation of the diagram in Fig. 16.     

Cytoarchitectonic pattern of the hypothalamus in the cobra,Naja naja

Summary The distribution and cytoarchitectonic pattern of the magno- and parvocellular hypothalamic nuclei of the cobra, Naja naja, are described at the light-microscopic level. With respect to their tinctorial affinity to paraldehyde fuchsin (AF) as a representative of the Gomori-type of stains, the magnocellular neurons belong to the AF-positive and the parvocellular neurons to the AF-negative elements. In addition to the supraoptic and paraventricular nuclei proper, two accessory aggregations of magnocellular neurons, the nucleus retrochiasmaticus and nucleus circularis, can be identified. Although in a peculiar location, they may be regarded as subunits of the supraopticoparaventricular neurosecretory complex. As many as 22 AF-negative nuclear areas are identified in the hypothalamus of the cobra. The nucleus periventricularis hypothalami of earlier authors is subdivided into several circumscribed neuronal complexes. The nucleus arcuatus, nucleus hypothalamicus lateralis and nucleus lateralis recessus infundibuli are well developed. An attempt is made to interpret the significance of these nuclei on a comparative and phylogenetic basis.On leave from the Department of Zoology, Nagpur University, Nagpur, India     

The development of the compound eyes ofPenaeus duorarum (Crustacea: Decapoda) with remarks on the nervous system

Summary The development of the compound eyes and nervous system of the penaeid shrimp,Penaeus duorarum, from the first nauplius to the first postlarva, has been studied. The first anlage of the compound eyes is a pair of optic discs on the front of the animal. These increase in size through cell-division until the second protozoea stage, where the eye-stalks appear with ommatidia and optic neuropiles developed. The original neuroectoderm of the optic discs is retained in the shape of a proliferation zone throughout the life of the animal. From the optic discs, develop the ommatidia, the lamina ganglionaris, and the medulla externa. The medullae interna and terminalis develop from cells coming from the brain anlage. From the second protozoea and onwards, the development is less rapid. The final shape of the adult eye is reached during the postlarval stages and includes the appearance of a few more pigments and a perfecting of several features. A scheme for the development of crustacean compound eyes is laid down. Further, the medulla externa of the Malacostraca and the single medulla of non-malacostracan crustaceans are homologized.The continuous growth of the nervous system is traced in the development of the neuropile. The appearance of glomeruli structures is reported, as are also, to some extent, neurosecretory organs. The development of the SPX-organ conforms to that of other decapods.For the sake of simplicity, the findings reported below in Results are grouped under two headings, namely the eye-stalk and the nervous system. Under the eye-stalk will be described both the structures coming from the optic discs comprising the ommatidia, the lamina ganglionaris, and the medulla externa, and the contributions from the nervous system comprising the medulla terminalis and medulla interna. Under the nervous system will be described the rest of the nervous system. The term anlage of the compound eyes is the same as the optic discs and denotes all contributions from this area in the early larva.     

The Releaser Effects of the Tergal Gland Secretion of Queen Honeybees (Apis mellifera)

Queen honeybees are attractive to their workers, due partially to the pheromonal bouquet they secrete. Queen mandibular gland pheromone is a powerful attractant to worker honeybees but it is not solely responsible for eliciting retinue behavior. The attractiveness of virgin queen tergal gland secretions and queen mandibular pheromone to worker honeybees was tested using a retinue bioassay. The number of workers attending the treated pseudoqueen lures was videorecorded in order to allow for the quantification of attractiveness. Queen mandibular gland secretions were more attractive than tergal gland secretions (P 0.008), and both queen tergal gland secretions (P 0.0002) and mandibular gland secretions (P 0.0001) were significantly more attractive than the control treatment. This laboratory bioassay indicates that queen tergal gland secretions have a releaser effect that evokes retinue behavior from worker honeybees.     

Intra- and extraganglionic nerve endings formed by neurosecretory cells of the cerebral ganglion of the earthworm (Lumbricus terrestris L.)

Summary In the cerebral (= supraesophageal, suprapharyngeal) ganglion of the earthworm, a number of neurosecretory Gomori-positive perikarya are bipolar; others are unipolar, or multipolar. Some of the neurosecretory cell processes project centrally into a fibrous zone; peripheral processes enter small nerves which leave the dorsocaudal aspect of the ganglion.In the central fibrous zone, the neurosecretory fibers form varicose Gomoripositive terminals. Here, also zinc-iodine-osmium (ZIO)-positive fibers and monoamine fluorescent fibers are found. With the electron microscope, nerve terminals containing synaptic vesicles and either large neurosecretory peptidergic granular vesicles (diameter more than 1500 Å), or smaller granular vesicles (diameter about 1300 Å, or 900 Å) are observed. These axon endings mainly form axo-dendritic synapses. Peptidergic profiles are both pre- and postsynaptic. Some of the extraganglionic peptidergic fibers appear to terminate around vessels, but most of them form terminals on the visceral muscle cells which surround the ganglion.We think that the central neurosecretory processes communicate with the fibers of the synaptic zone of the ganglion. The peripheral neurosecretory peptidergic fibers are supposed to form a primitive neurohemal area and/or to function as vasomotor nerves. The fibers innervating the visceral muscle cells may represent vegetative nerves.     

Immunohistochemical localization of dopamine in the brain of the insect Locusta migratoria migratorioides in comparison with the catecholamine distribution determined by the histofluorescence technique

Summary As part of a follow-up study to our previous investigation of the catecholaminergic neurosecretory cells in the brain of adult female locusts (Locusta migratoria migratorioides) by means of the formaldehyde-induced fluorescence method, we have attempted to specify the identity of the amines present in these cells by an immunohistological technique. Using a recently developed anti-dopamine serum, we have demonstrated that the majority of the cate cholaminergic median neurosecretory cells contain dopamine. Moreover, dopamine is present in some cell bodies of other zones of the brain, i.e. the median subocellar neurosecretory cells, perikarya in external areas of the protocerebrum, below the calyces, around the pedunculus, in the optic lobes (between the lobula and the medulla, between the medulla and the lamina), and in external zones of the tritocerebrum. Among the structured neuropils, which were particularly fluorescent in the formaldehyde-induced fluorescence method, only the pedunculus, the posterior part of the central body, the external zones of the - and lobes and the proximal part of the lamina contain little dopamine.     

Ultrastructure and potential significance of cerebral light-refracting bodies of Stenostomum virginianum (Turbellaria,Catenulida)

Summary Light-refracting bodies, possibly photoreceptors, occurring in the posterior lobes of the brain are considered characteristic for most species of catenulid turbellarians of the freshwater genus Stenostomum. In S. virginianum, each of two light-refracting bodies consists of a single cup-shaped granule (3–4 m) situated in the perikaryon of a single nerve cell (the clear vesicle of earlier papers). TEM reveals each granule as an enlarged and folded mitochondrion with a dense matrix inclusion of undetermined composition. Cristae are well-developed and there is a dense granular extramito-chondrial layer of uniform thickness (100 nm) along the posteromedial surface of the mitochondrion. The perikaryon is packed with ribosomes, -glycogen granules and 60–100 nm dense-cored vesicles. A neurite extends from the perikaryon into the neuropile of the brain. Experimental data indicate an absence of phototaxis and photokinesis and an absence of ultrastructural modifications with light- and dark-adaptation. An ultrastructural comparison is made of the light-refracting bodies of S. virginianum with those of a second species. Hypotheses regarding the role of light-refracting bodies as photoperiodic receptors and/or specialized neurosecretory cells are advanced.This study was supported in part by a National Science Foundation grant DEB-7823395 and a Clemson University Faculty Research Grant to E. Ruppert     

Immunocytochemical localization of CCAP,a novel crustacean cardioactive peptide,in the nervous system of the shore crab,Carcinus maenas L.

Summary Polyclonal antibodies were raised in rabbits against synthetic crustacean cardioactive peptide (CCAP) conjugated to bovine thyroglobulin, and were used to map CCAP-immunoreactive structures in the central nervous system of Carcinus maenas. As expected, the neurohemal pericardial organs (PO) displayed abundant immunoreactivity in nerve fibers and terminals. In addition, immunoreactive neurons were demonstrated in other parts of the nervous system. At least some of them do not appear to terminate in neurohemal structures and may have a non-endocrine, as yet unknown function. Immunoreactive perikarya with a diameter of 25–30 m occur in the brain. They project into the optic and antennary neuropil, and into the eyestalk. One cell was found in the medulla terminalis of the eyestalk and in the connective ganglion, respectively. From the latter, axonal branches could be traced into the brain and the thoracic ganglia (TG). In the TG, small-diameter perikarya give rise to extensive networks of varicose fibers. Some of the perikarya occur in a characteristic paired arrangement with larger CCAP-immunoreactive somata (diameter 40–50 m). These pairs of one small and one large cell occur in all mouthpart and leg segments of the TG, except the abdominal ganglia (AG), where only large cells were found. The main projections of the large neurons comprise one or more fibers in each of the seven segmental nerves (SN), leading to neurosecretory terminals in the PO. The fibers in the SN are joined by branches of an ascending axonal tract from the large perikarya in the AG. The large-type perikarya are considered to be the principal source of CCAP in the PO. The optic ganglia in the eyestalk, except the medulla terminalis, the neurohemal sinus gland and the stomatogastric nervous system are devoid of CCAP-immunoreactivity.In axon terminals of the PO, CCAP is not colocalized with other PO-neuropeptides, i.e. proctolin-, FMRFamide-like, and Leu-enkephalin-like immunoreactive materials. Electron-microscopic immunocytochemistry revealed a distinct CCAP-containing granule type in specific axon profiles and terminals in the PO.The architecture of CCAP-immunoreactive neurons is discussed with respect to previous morphological studies on the origin and pathways of fibers terminating in the PO.Dedicated to Professor K.E. Wohlfarth-Bottermann, Bonn, on the occasion of his 65^th birthday     

Structural comparison of the genes of two HLA-DR supertypic groups: The loci encoding DRw52 and DRw53 are not truly allelic

The organization and sequence of the HLA-DR chain genes are compared in the two supertypic groups, DRw52 and DRw53, which together account for more than 80% of HLA-DR alleles. From the structural data, we conclude that these two groups represent distinct lineages which have followed different patterns of evolution. The fine structure of the chain locus encoding the DRw53 specificity corresponds most closely to the DR II pseudogene in the DRw52 haplotypes. Concomitantly, the DR I locus in DRw53 haplotypes is more closely related to both of the two expressed DR loci of theDRw5 haplotypes (DR I and DR III). These two loci are the result of a recent duplication. This leads to the proposal that both expressed DR chain genes in the DRw52 haplotypes (DR I and DR III) are derived from a single precursor locus, while the two loci expressed in the DRw53 haplotypes are derived from distinct ancestral loci. The genes encoding DRw52 and DRw53 are therefore not true alleles of the same original locus. A scheme is proposed that accounts for the evolution of DR specificities within the DRw52 and DRw53 groups of haplotypes. It is evident that the differentHLA-DR alleles are not structurally equidistant and that one must take into consideration different degrees of heterozygosity or mismatch among the DR alleles.     

Hormonabhängige Enzyraverteilung in Geweben

Zusammenfassung Bei 46 männlichen kastrierten, z.T. zusätzlich adrenalektomierten Ratten wurden nach unterschiedlicher Behandlung mit Thyroxin die spezifischen Aktivitäten der alkalischen und sauren Phosphatase in Rinde und Außenstreifen der Niere bestimmt. Die Untersuchungen wurden durch histochemische Enzymdarstellungen ergänzt. Die Resultate stellen sich wie folgt dar:Eine Aktivitätssteigerung der alkalischen Nierenphosphatase kann mit einer einmaligen Gabe von 60 g Thyroxin s.c./100 g KG erzielt werden. Nur der Außenstreifen (gestreckte Hauptstücke) reagiert. Im Mittel wird eine maximale Aktivitätszunahme 72 Std nach der Thyroxingabe erreicht. Der Unterschied gegenüber der Kontrolle beträgt dann 10,9 MKH, der Zuwachs mithin 90%. Zusätzlich adrenalektomierte Tiere reagieren in ähnlicher Weise.Eine weniger ausgeprägte Aktivitätszunahme findet sich auch nach 6tägiger Behandlung mit täglich 15 g Thyroxin: 100 g KG. Der Zuwachs im Außenstreifen beträgt im Mittel 4,7 MKH = 39%.Die saure Phosphatase wurde nach einmaliger Thyroxingabe in Rinde und Außenstreifen erniedrigt gefunden.

---

Hormone dependent enzyme distribution in animal tissueXVII. The influence of thyroxine on the phosphomonoesterases of the rat kidney

Summary In 46 male castrated rats, some of them being additionally adrenalektomized, the specific activities of the alkaline and acid kidney phosphatase in cortex and outer stripe of the medulla were determined quantitatively following different treatment with thyroxine. The investigations were supplied by histochemical demonstration of enzymes. The results are as follows:An increase of alkaline phosphatase in the kidney can be reliably reproduced by a single dose of 60 g thyroxine/100 g b.w. Only the outer stripe of the medulla (= straight portion of proximal tubules) reacts. In the medium a maximal increase is reached 72 hours after application of thyroxine. The difference against the control amounts 10.9 MKH, i.e. 90%.Additionally adrenalektomized rats react in a similar manner.A less pronounced increase of enzyme activity is found following a daily repeated treatment with 15 g thyroxine/100 g b.w. over a period of six days. The increase in the outer stripe of the medulla amounts in the medium 4.7 MKH = 39 %.After a single dose of thyroxine, the acid phosphatase was found decreased in both cortex and outer stripe of the medulla.

---

---

Mit Unterstützung durch die Deutsche Forschungsgemeinschaft, Bad Godesberg.

---

Wesentliche Teile der vorliegenden Arbeit wurden von Herrn Rimek als Dissertation der Medizinischen Fakultät der Universität Freiburg i. Br. vorgelegt.     

Synaptic organization of a multifunctional interneuron in the central nervous system of Helix pomatia L.

Summary The morphology, axonal arborization and ultrastructure of synaptic connections of the V21 giant neuron in the visceral ganglion of the snail Helix pomatia has been investigated following intracellular labelling with horseradish peroxidase. The V21 neuron establishes several afferent and efferent axo-axonic connections, mainly along the first half of the primary axon. Collaterals of 200–300 m length originate from the primary axon, which shows further arborization, and both afferent and efferent synaptic contacts are formed on these fine axon profiles. Afferent and efferent contacts of the cell occur within very short distances of a few micrometers. On the basis of ultrastructure and vesicle and granule content, several afferent terminals can be distinguished on V21 labelled axon profiles. The majority of these afferent terminals resembles peptidergic-(neurosecretory)-like terminals. This finding supports the possible transmitter role of neuropeptides in the central nervous system of gastropods. Our results are consistent with and provide morphological evidence for recent electrophysiological observations suggesting that, in addition to integrating input, the V21 neuron functions as an interneuron in Helix central nervous system.     

Weitere elektronenmikroskopische Untersuchungen am kaudalen neurosekretorischen System von Fischen

Zusammenfassung Das kaudale neurosekretorische System von Cyprinus carpio und Channa argus Cantor wurde elektronenmikroskopisch untersucht. Während in den Golgi-Bläschen von Channa nur eine Art von Elementargranula entsteht, kommen bei Cyprinus zwei Granulatypen verschiedener Größe und Elektronendichte (- und ß-Granula) vor, jedoch nie in ein und derselben neurosekretorischen Zelle zusammen. Bei Cyprinus werden Elementargranula verschiedener Elektronendichte aus der Zelle in die perivaskulären Räume abgegeben.Außer Elementargranula wurden große Tropfen mit verschiedener Struktur in den Perikaryen und in den Basen der Zell-Fortsätze beobachtet. Ein Teil der Tropfen ist durch Verschmelzung der Elementargranula entstanden, die meisten jedoch entsprechen Lysosomen, Myelinfiguren, multivesicular bodies usw.Die neurosekretorischen Zellen sind durch zahlreiche axo-somatische und axo-axonale sowie axo-dendritische Synapsen mit nichtsekretorischen Nervenzellen verbunden. — Da die Wandungen der endozellulären Kapillaren ohne Zwischenschaltung einer gliösen Schranke die Zellmembran der Perikaryen berühren, wird ein unmittelbarer Stoffaustausch zwischen neurosekretorischen Zellen und Blut vermutet.

---

Summary The caudal neurosecretory system of Cyprinus carpio and Channa argus Cantor was studied electron microscopically. In the secretory nerve cells of both species the elementary granules are formed in the Golgi complex. In the neurosecretory system of Cyprinus carpio two kinds of neurosecretory cells can be differentiated according to the size and the electron density of their granules (- and ß-granule). Grnules of both types cannot be found in the same neurosecretory cell. In Cyprinus the elementary granules of various density are released into the pericapillary space.Apart from the elementary granules droplets with various structures in the perikarya and bases of the processes were recognized. A part of these droplets consists of accumulated elementary granules, the majority however belongs to inclusions like lysosomes, myelinfigures, multivesicular bodies and so on.The neurosecretory elements are connected with non-secreting nerve cells by numerous axo-somatic, axo-axonic and axo-dendritic synapses. In Channa argus the perikarya of neurosecretory cells are in close contact with the basement membrane of the capillary, i.e. there is no glial barrier between the neurosecretory cell and the blood vessel.

---

---

Frau Professor Dr. B. Scharrer zum 60. Geburtstag gewidmet.     

Iodination activity in Eisenia foetida (Annelida,Oligochaeta)

Summary In the nervous system of the earthworm Eisenia foetida, we have previously found thyroglobulin-like immunoreactive neurons. In the present study iodination activity was investigated by injecting worms or incubating them in vivo with radioiodine; animals treated with methimazole (MMI), an inhibitor of peroxidase-catalyzed iodination, served as controls. Radioiodinated proteins were identified in soluble extracts from ¹²⁵I-incubated animals; the sedimentation pattern of soluble proteins from cephalic segments showed a peak of radioactivity in the 3–4 S region. In animals pretreated with 10^-3 M MMI for 48 h, ¹²⁵I-incorporation into soluble proteins from cephalic segments was drastically reduced. Light-microscopic autoradiographic studies showed silver-grains selectively concentrated in the brain and ventral nerve cord of ¹²⁵I-injected animals. The highest grain-density occurred in the cerebral ganglion beginning 5 min after tracer injection; the reaction product was mainly distributed between the neurosecretory cells and neuropile fibres in the zone of the presumptive plexiform neurohaemal complex. In MMI-pretreated animals the reaction product was not visible in either cerebral ganglion or ventral nerve cord. Of interest, the setae appeared consistently positive with or without MMI treatment. These observations indicate that protein iodination involving peroxidase occurs in the nervous system of earthworms and suggest that iodination mechanisms, other than peroxidase-catalyzed, may be operating in some scleroprotein structures such as setae.     

Characterization of buccal motor programs elicited by a cholinergic agonist applied to the cerebral ganglion of Aplysia californica

Applying the non-hydrolyzable cholinergic agonist carbachol (CCh) to the cerebral ganglion of Aplysia elicits sustained, regular bursts of activity in the buccal ganglia resembling those seen during biting. The threshold for bursting is 10^2–4 M. Bursting begins after a 2 to 5 min delay. The burst frequency increases over the first 5 bursts, reaching a plateau value of 3 per minute. Bursting is maintained for over 10 min. Some of the effects of CCh may be attributed to its ability to depolarize and fire CBI-2, a command-like neuron in the cerebral ganglion that initiates biting. CBI-2 is also depolarized by ACh, and by stimulating peripheral sensory nerves. Excitation of CBI-2 caused by carbachol is partially blocked by the muscarinic antagonist atropine. We examined whether CCh-induced bursting is modified in ganglia taken from Aplysia that previously experienced treatments inhibiting feeding, such as satiation, head shock contingent or non-contingent with food, and training animals with an inedible food. No treatment consistently and repeatedly affected the latency, the peak burst period, the length of time that bursting was maintained, or the threshold CCh concentration for eliciting bursting. However, there was a decrease in the rate of the buildup of the buccal ganglion program in previously satiated animals.     

An approach for parameter estimation of biotechnological processes

An approach for parameter estimators design of biotechnological processes (BTP) is presented in case of lack of real time information about state variables. It is based on general reaction rate models and measurements of at least one reaction rate. A general parameter estimator of BTP is designed with the help of which specific rate estimators are synthesized. Stability and convergence of an estimator of specific growth rate for a class of aerobic batch processes are proved. Its effectiveness is illustrated by simulation results. The proposed on-line parameter estimation approach can be used for design of BTP on-line variable estimation algorithms (variable observers of BTP).List of Symbols X, S, P g/l biomass, substrate and product concentrations - C g/l oxygen concentration in the culture broth - C _sg/l saturation concentration of oxygen in the culture broth - C _in, C_outg/l oxygen concentrations in the input air flow and in the outlet gasphase - F _in, F_outl/h the input air flow in the fermenter and output air flow - OUR g/(lh) oxygen consumption rate - OUR _mg/(lh) measured values of OUR - V l volume - , , l/h specific growth, consumption and synthesis rates - K _La(o) l/h specific volumetric mass transfer coefficient - D l/h dilution rate - R _X, R_S, R_Pg/(lh) biomass growth, substrate consumption and product synthesis rates - K _b matrix of yield coefficients - H_b(), H() matrices of known functions of - H(R) matrix of known functions of R - and gain matrices - a vector of the state variables - () a reactions rates vector, describing qualitative relations among the components - R() a reactions rates vector, describing qualitative and quantitative relations among the components - F a feed rates vector - Q a gaseous outflow rates vector - _b () a vector of unknown functions of - ₁() a vector of functions - (t) a vector of unknown time-varying parameters - ₂(, ) an auxiliary vector-function of and - Y _X/S, Y_X/C, Y_X/P substrate, oxygen and product yield coefficients - b maintenence coefficient - k _i(i=1...6) kinetic coefficients - C _i(i=1,2) design parameters estimate     

An electron microscopical investigation of the follicle gland (cerebral gland) and of some neurosecretory cells in the lateral lobe of the cerebral ganglion of the pulmonate gastropod Lymnaea stagnalis L.

Summary The ultrastructure of the follicle gland and of some Gomori-positive neurosecretory cells in the lateral lobes of the cerebral ganglia of Lymnaea stagnalis is described.The follicle wall consists of epithelial cells containing secretion granules, and of processes of lateral lobe nerve cells bearing cilia, including the processes of bipolar neurosecretory B-cells. The ultrastructure of the follicle of Ancylus fluviatilis and Planorbarius corneus appeared to be very similar to that of L. stagnalis. From the ultrastructure of the follicle wall and lumen, no conclusions can be drawn as to its formerly proposed possible functions (sense organ, endocrine organ, neuroendocrine organ). Of three neurosecretory cell types (B-cells, canopy cells, droplet cells), the positivity to chrome-haematoxylin and paraldehyde-fuchsin appeared to be due to elementary granules of different size and appearance. This finding supports in a particular sense the view that results obtained with the neurosecretory stains, should be judged carefully. Also in ordinary neurons elementary granules can be found, at times in great numbers. Several types of elementary granules (at least 6) could be distinguished. Their character and function (neurohormones, neurotransmitters) are not clear.     

Histological and histochemical observations on the neurosecretory cells in the diencephalon of Chthonerpeton indistinctum and Ichthyophis paucisulcus (Gymnophiona,Amphibia)

Summary In the diencephalon of two species of Gymnophiona (Amphibia) two neurosecretory nuclei were examined with histological (Alcian Blue, Aldehyde Fuchsin, Brookes Trichrome stain) and enzyme histochemical techniques (acid phosphatase, -naphthyl acetate esterase, acetylcholinesterase (AChE)). In the preoptic nucleus two categories of secretory neurons were distinguished: large and medium sized neurons. The perikarya of both cell types contain very little neurosecretory material. The Alcian Blue method stained the medium sized neurons faintly but selectively. The tractus praeopticohypophyseus is marked by the presence of Herring bodies, which, however, are relatively scarce. The neurohypophysis, in contrast, contains large amounts of neurosecretory material. Both cell types of the preoptic nucleus are characterized by their very strong AChE and -naphthylacetate esterase activity. The AChE also marks the tractus praeoptico-hypophyseus. In the large neurons acid phosphatase is present around the nucleus; in the medium sized neurons this enzyme is concentrated close to the origin of the axon. In the dorso-caudal hypothalamus a small group of neurons is stained with Alcian-Blue. These neurons, which also contain AChE, are located immediately under the ependyma which seems to be specialized in this region.The financial support of the Deutsche Forschungsgemeinschaft is gratefully acknowledged (We 380/5)     

The frontal ganglion of Periplaneta americana L. (insecta)

Summary The frontal ganglion, part of the stomatogastric nervous system, contains about 60 to 80 neurons, 25 to 30 m in diameter. A well developed Golgi system, producing dense-core vesicles, lysosomes, multivesicular bodies and dense bodies are abundant. Glia elements are sparsely distributed. Many nerve fibres contain granules of different size and electron density. Five groups of fibres can be distinguished: Fibres with granules of about 200 nm (type A), fibres with granules of about 160 to 170 nm (type B), fibres with granules of about 80 to 100 nm (type C) and those with synaptic vesicles of 50 nm (type D) respectively. A fifth very small type contains neither vesicles nor granules. Special attention was paid to synaptic contacts. The divergent dyad seems to be the main type in the frontal ganglion. Frequently, neurosecretory endings are observed in presynaptic position. Immunocytochemical staining of neurosecretory material closely corresponds to the distribution of type A fibres, as observed electron microscopically. Immunoelectrophoresis of extracts from frontal ganglia with polyspecific anti-neurosecretion-serum reveals a single precipitation line, indicating that the immunocytochemical localization of neurosecretory material is due to reaction with a specific as well as a crossreagent antibody.Supported by the Ministerium für Wissenschaft und Technik der DDRThe authors wish to thank Mrs. B. Cosack and Mrs. A. Schmidt for excellent technical assistance     

Serotonin-like immunoreactivity in the ventral nerve cord of the Colorado potato beetle,Leptinotarsa decemlineata: Identification of five different neuron classes

Summary In an immunohistochemical study of the ventral nerve cord of L. decemlineata, five distinct neuron categories were distinguished: 1) Two paired segmental twin interneurons occur in each ganglion or neuromere; their axons distribute processes over almost the entire nerve cord and run to the cerebral ganglion complex. In contrast, other axons are distributed locally. 2) Four large frontal neurosecretory neurons occur in the suboesophageal ganglion (SOG), two of which have axons that run into the mandibular nerves to form a neurohemal plexus on the surface of cerebral nerves. 3) A pair of large caudal neurons occur in the terminal ganglion and innervate the hindgut. 4) Local miniature interneurons occur in the SOG. 5) Terminal neurons are present in the last abdominal ganglion. Segmental twin interneurons appear to be grouped into 3 functional units spanning several ganglia. Their axons run to specific projection areas, which separate the functional units, and which mark the externally visible separation of condensed ganglion complexes. A possible role of the most caudal functional unit might be the synaptic control of caudal neurons innervating the hindgut.     

Purification and properties of a novel β-galactosidase or exo-(1 → 4)-β-d-galactanase from the cotyledons of germinated Lupinus angustifolius L. seeds

The main polysaccharide component of the thickened cell walls in the storage parenchyma of Lupinus angustifolius L. cotyledons is a linear (1 4)--linked d-galactan, which is mobilised after germination (L.A. Crawshaw and J.S.G Reid, 1984, Planta 160, 449–454). The isolation from the germinated cotyledons of a -d-galactosidase or exo-(1 4)--d-galactanase with a high specificity for the lupin galactan is described. The enzyme, purified using diethylaminoethyl-cellulose, carboxymethyl-cellulose and affinity chromatography on lactose-agarose, gave two bands (major 60 kDa, minor 45 kDa) on sodium dodecyl sulphate-gel electrophoresis, and two similar bands on isoelectric focusing (major, pI 7.0, minor pI 6.7, both apparently possessing enzyme activity). The minor component cross-reacted with an antiserum raised against, and affinity-purified on, the major band. Both components had a common N-terminal sequence. The minor component was probably a degradation product of the major one. The enzyme had limited -galactosidase action, catalysing the hydrolysis of p-nitrophenyl--d-galactopyranoside and (1 4)- and (1 6)--linked galactobioses. Lactose [-d-galactopyranosyl-(1 4)-d-glucose] was hydrolysed only very slowly and methyl--d-galactopyranoside not at all. Lupin galactan was hydrolysed rapidly and extensively to galactose, whereas other cell-wall polysaccharides (xyloglucan and arabinogalactan) with terminal non-reducing -d-galactopyranosyl residues were not substrates. A linear (1 4)--linked galactopentaose was hydrolysed efficiently to the tetraose plus galactose, but further sequential removals of galactose to give the tetraose and lower homologues occurred more slowly. Galactose, -galactonolactone and Cu⁺² were inhibitory. No endo--d-galactanase activity was detected in lupin cotyledonary extracts, whereas exo-galactanase activity varied pari passu with galactan mobilisation. Exo-galactanase protein was detected, by Western immunoblotting of cotyledon extracts, just before the activity could be assayed and then increased and decreased in step with the enzyme activity. The exo-galactanase is clearly a key enzyme in galactan mobilisation and may be the sole activity involved in depolymerising the dominant (1 4)--galactan component of the cell wall.Abbreviations CM carboxymethyl - DEAE diethylaminoethyl - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - TLC thin-layer chromatography We wish to thank CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) for the award of a studentship to M.S. Buckeridge, and the Government of São Paulo State, Brazil for granting him leave of absence. We are grateful to Dr. Amanda Heyller (Unilever Research Laboratory, Colworth House, Bedford, UK) for N-terminal sequence determinations, to Dr. Stuart Wilson (Stirling) for preparing gelatin SDS-gels and to Cristina Fanutti (Stirling) for purifying the xyloglucan oligosaccharide.