Studies on the neurosecretory system and retrocerebral endocrine glands of Nezara viridula Linn. (Heteroptera: Pentatomidae)

The neurosecretory system and retrocerebral endocrine glands of Nezara viridula Linn. have been described on the basis of in situ preparations and histological sections employing the paraldehyde fuchsin (PF) and performic acid-victoria blue (PAVB) techniques. In the brain of N. viridula, there are two medial groups–each consisting of five neurosecretory cells which belong to A-type. The lateral neurosecretory cells are absent. The axons of the two groups of medial neurosecretory cells (MNC) compose the two bundles of neurosecretory pathways (NSP) that decussate in the anterodorsal part of the protocerebrum. The two pathways, after the cross-over, run deep into the protocerebrum and deutocerebrum and emerge as NCC-I from the tritocerebrum. The nervi corporis cardiaci-I (NCC-I) of each side which are heavily loaded with NSM terminate in the aorta wall. Thus, the neurosecretory material (NSM), elaborated in the medial neurosecretory cells of the brain, is stored in the aortic wall and nervi corporis cardiaci-I (NCC-I). The NCC-II are very short nerves that originate from the tritocerebrum and terminate in the corpora cardiaca (CC) of their side. Below the aorta, but dorsal to the oesophagus, lie two oval or spherical corpora cardiaca. A corpus allatum (CA) lies posterior to the corpora cardiaca (CC). The corpora cardiaca do not contain NSM; only the intrinsic secretion of their cells has been occasionally observed which stains orange or green with PF staining method. The corpus allatum sometimes exhibits PF positive granules of cerebral origin. A new connection between the corpus allatum and aorta has been recorded. The suboesophageal ganglion contains two neurosecretory cells of A-type which, in structure and staining behaviour, are similar to the medial neurosecretory cells of the brain. The course and termination of axons of suboesophageal ganglion neurosecretory cells, and the storage organ for the secretion of these cells have been reported. It is suggested that the aortic wall and NCC-I axons function as neurohaemal organ for cerebral and suboesophageal secretions.     

Neurosecretory cells in the optic lobe of two aquatic coleopteran species,Dineutes indicus aube and Cybister rugulosus redt

In each optic lobe and optic peduncle of two aquatic beetles viz. Dineutes indicus and Cybister rugulosus the neurosecretory cells are observed with the help of various histochemical techniques. These cells are arranged to form a discrete group. A group in the optic lobe of both species contains about 25 to 30 neurosecretory cells. On the basis of staining properties the neurosecretory cells are classified into A and B types. These cells stain with chrome haematoxylin-phloxine and paraldehyde fuchsin, but do not stain with azan. Histochemically, the neurosecretory material is positive for proteins and shows a negative reaction for 1,2-glycols. The cells show variations in RNA contents in correlation with the state of secretory activity. Axons of the neurosecretory cell group of the optic lobe are observed directed to the optic peduncle. The axonal tract from neurosecretory cells in the optic peduncle runs towards the lateral margin of the brain.     

The distribution of the urban mosquitoes, Culex pipiens pipiens f. molestus (Diptera: Culicidae) in Petrozavodsk City

The distribution of the urban mosquitoes, Culex pipiens pipiens f. molestus in Petrozavodsk city was studied for the first time. Immature mosquito stages were found in 21 of 68 basements flooded with polluted waters, the occurrence of the mosquitoes varied in different districts of the sity. The mean siphonal index of the mosquito larvae from 6 underground populations varied from 3.8 up to 4.2, i. e. within the range typical for C. p. p. f. molestus.     

Molecular genetic methods for the identification of the urban mosquito Culex pipiens pipiens F. molestus (Diptera, Culicidae)

Molecular genetic methods for the identification of two ecotypes, or forms (pipiens and molestus) of the Culex pipiens pipiens mosquitoes, which are known as active bloodsuckers and vectors of various agents of diseases, are proposed. For the DNA analysis, two populations of the urban mosquitoes (the molestus ecotype) from St. Petersburg and Moscow and two populations from the overground reservoirs in the Leningrad Province and neighboring areas of Moscow (the pipiens ecotype) have been studied. These ecotypes differ by six transitions among 247 nucleotide sequences of 3' part of the cytochrome oxidase subunit I (COI) gene of mitochondrial DNA (mtDNA), by the mtDNA fragments resulted from the restriction analysis and by the lengths of second internal transcribed spacer (ITS2) sequences in the ribosomal DNA.     

Histology, histochemistry, and ultrastructure of neurosecretory cells in the optic lobe of the cockroach, Periplaneta americana

In the region of the distal optic chiasma of each optic lobe of Periplaneta americana, there is a group of about 120 monopolar neurosecretory cells. These cells do not stain with paraldehyde fuchsin but remain acidophilic after oxidation. They stain red or sometimes indigo with the azan technique. Histochemically, the neurosecretory material is positive for protein and the amino acids tryptophan and arginine but negative for 1, 2-glycols and strongly acidic groups. At the ultrastructural level, the cytoplasm of the cells contain many elementary neurosecretory granules 100 to 170 nm in dia. The cells also contain well-developed Golgi bodies and endoplasmic retieulum. The axons from these cells run toward the interior of the optic lobe. In this region, axons containing dense granules (mean diameter 70 nm) and synaptic vesicles synapse onto the axons from the neurosecretory cells. The neurosecretory axons then cross over to the anterior side of the optic lobe and run towards the brain. The function of these neurosecretory cells is unknown, but they may be involved with photoperiodically controlled activity rhythms.     

Cultivation of the mosquito Culex pipiens pipiens f. molestus (Diptera, Culicidae) without blood feeding

Optimal photoperiod and temperature conditions for rearing of the mosquito Culex pipiens pipiens f. molestus without blood feeding (through autogenous eggs) were experimentally substantiated. This method may be used in order to obtain relatively small quantities of mosquitoes or for preservation of a laboratory colony. Optimal conditions of rearing are the short-day photoperiod (12 h of light per day) at 20 degrees C, when 87 % of the females produce autogenous eggs rafts consisting of 52 eggs on the average. It should be emphasized that larval food must be adequate and abundant, because the quality and quantity of it has a pronounced effect on the autogeny rate.     

Immunofluorescent localization of a substance immunologically related to insulin in the protocerebral neurosecretory cells of the European corn borer

Summary A substance immunologically related to mammalian insulin was demonstrated by immunofluorescence in one of five types of neurosecretory cells in the protocerebrum of the European corn borer, Ostrinia nubilalis. This A₂ cell type contains a secretory material stainable with paraldehyde fuchsin.E.R.A. CNRS n° 850     

Ultrastructure of neurosecretory cells in the pars intercerebralis of Rhodnius prolixus (Hemiptera).

Six neuron types are distinguished in the pars intercerebralis of the starved fifth instar of Rhodnius prolixus. All neuron types contain electron dense secretory granules derived from Golgi complexes which are of characteristic size and morphology in each type. The neuron types are not thought to represent stages in a secretory cycle. The variety of neuron types described is related to that revealed by staining sections of the same cells with paraldehyde fuchsin. Active synthesis of neurosecretory granules continues throughout starvation and the lysosomal system appears to be involved in the continual degradation of secretory granules. Some of the variations in granule morphology observed may be a consequence of granule fusion and the importance of cytoplasmic events in the development of neurosecretory granules is discussed.     

Staining properties of aldehyde fuchsin analogs.

This investigation was designed to clarify the role of the aldehyde component of aldehyde fuchsin in its staining reactions. Several aldehyde fuchsin analogs were prepared by using different aldehydes. The staining quality of these analogs and pararosaniline-HCl was compared with that of aldehyde fuchsin prepared with paraldehyde in the usual way. The major findings of this investigation include: 1) Aldehyde fuchsin staining of nonoxidized pancreatic B cells requires a stain prepared with either paraldehyde or acetaldehyde. 2) An aldehyde moiety is required for aldehyde fuchsin staining of strong tissue anions. 3) Staining of elastic tissue with aldehyde fuchsin analogs resembles staining of strong tissue anions more than staining of nonoxidized pancreatic B cells. Possible reaction mechanisms of aldehyde fuchsin with tissue substrates are discussed.     

The role of paraldehyde in the rapid preparation of aldehyde fuchsin

Preparation of aldehyde fuchsin normally requires ripening for 3 to 5 days. By using a 5-fold excess of paraldehyde a fully potent aldehyde fuchsin can be prepared in 24 hr at room temperature. Aldehyde fuchsin prepared by both normal and accelerated ripening afforded comparable results, including selective staining of unoxidized pancreatic B cells. Dried aldehyde fuchsin prepared form pararosaniline and reconstituted in acid alcohol has spectrophotometric properties different form the ripened strain. Reconstituted aldehyde fuchsin stains unoxidized B cells adequately only if staining time is extended. Excess paraldehyde added to reconstituted aldehyde fuchsin retards decomposition but does not produce a normal stain by spectrophotometric standards. Warming of aldehyde fuchsin solutions to accelerate ripening has been shown to produce deleterious effects and should be avoided.     

中国尖音库蚊复合组支序系统学的研究

本文应用支序系统学原理和方法,采用形态和生态性状,对尖音库蚊复合组的４个成员进行了支序系统学研究,提出在尖音库蚊复合组中,尖音库蚊是原始的类群,骚扰库蚊为较进化的类群,而致倦库蚊和淡色库蚊是最为进化的类群。     

Characterization of a new neurosecretory cell type containing gastrin-cholecystokinin-like peptide in the locust pars intercerebralis: ultrastructural and immunocytochemical studies,in situ and in vitro

A new neurosecretory cell type of the locust pars intercerebralis, immunolabelled with an antiserum against a vertebrate peptide related to gastrin-cholecystokinin (CCK-8(s)), was characterized both in situ and in primary cell cultures. Semithin sections of pars intercerebralis were first immunostained in order to identify neurosecretory cells containing CCK-like material and then examined by electron microscopy. The neurosecretory cells containing CCK-like material were paraldehyde fuchsin negative and were unequivocally identified in ultrathin sections adjacent to immunostained semithin sections. They exhibited neurosecretory vesicles of variable electron density, ranging in diameter from 150 to 250 nm. Immunogold labelled ultrathin sections adjacent to unlabelled ultrathin sections allowed for the unambiguous localization of CCK-like immunoreactive material over the neurosecretory vesicles of the cells containing CCK-like material. Immunoreactivity towards CCK-8(s)-like peptide could also be detected in pars intercerebralis neurosecretory neurons grown in vitro. The CCK-like positive neurons showed a multipolar morphology with fine processes radiating from the cell body. The positive cells had the same ultrastructural characteristics as the in situ CCK-like neurons. The pattern of neurite outgrowth on reactive CCK-like neurosecretory cells in vitro and the neuroanatomical pathway of the CCK-like immunoreactive neurosecretory cells in situ could be correlated. On the basis of their number, size and localization in the locust pars intercerebralis, it is possible that the CCK-like neurosecretory cells correspond to neurosecretory cell type C, which has not, to date, been identified at the ultrastructural level.     

Ultrastructural evidence for the existence of two types of neurosecretory cells in the abdominal ganglia of the chelicerate arthropod,Limulus polyphemus

Evidence suggesting the existence of two types of neurosecretory cells in each abdominal ganglion of Limulus polyphemus has been obtained by light and electron microscopy. After Helly fixation the two cell types are readily distinguished from other neurons by the Azan method, but they react weakly when stained by paraldehyde fuchsin. Type I cells are larger, more regular in shape, and found more anteriorly in each ganglion. They contain apparently cylindrical secretory granules, many dictyosomes, and numerous cytoplasmic vesicles. Type II cells produce spherical granules, contain fewer dictyosomes, have less conspicuous cytoplasmic vesiculation and possess more prominent parallel arrays of rough endoplasmic reticulum. Granules similar to those found in both cell types are present in the neuropile and certain nerves, but the specific pathways of the axons of these cells have not yet been determined.     

Polymorphism of mitochondrial DNA and infection with symbiotic cytoplasmic bacterium Wolbachia pipientis in mosquitoes of the Culex pipiens complex from Russia

A total of 208 mosquitoes of the Culex pipiens complex from 15 basement and terrestrial populations collected in different regions of the European part of Russia and Siberia were examined by genetic methods. Among these, two major mitotypes, M and P, were identified. These mitotypes differed by six substitutions in the 246-bp mitochondrial DNA cytochrome oxidase I gene fragment examined. Urban basement mosquito ecotype C. pipiens (form molestus) were characterized by the presence of mitotype M and infection with the endosymbiotic bacteria of the genus Wolbachia. Mosquitoes of the C. pipiens complex inhabiting opened biotopes harbored mitotype P, or its variety, mitotype P1, and were not infected with Wolbachia. Thus, in natural conditions marked linkage disequilibrium between cytoplasmic elements, mitochondrial DNA and Wolbachia, can be observed. Similarity of mitotypes in form molestus mosquito from different geographical localities favors the hypothesis on the common ancestry of urban mosquitoes.     

A histochemical study of the cells of the ventral cord ganglia of the horseshoe crab,Limulus polyphemus (L.)

Summary The ventral cord ganglia of the horseshoe crab, Limulus polyphemus, contains six distinct cell types: three appear to be ordinary neurons and three exhibit the staining affinities of neurosecretory cells.The presumed neurosecretory cells have been termed neurosecretory cell I (NSC I), NSC II and NSC III. NSC I cells contain a colloid-like inclusion which may occur as a single small vacuole or occupy more than one-half of the cell volume. Colloid inclusions occur with greater frequency toward the periphery, although small cells of similar staining affinity occur in cords extending to the fibrous core. The histochemical tests suggest that the cytoplasm is positive for proteins, but contains no strong acidic groups which may have been derived from S-S or S-H groups. The presence of carbohydrate is also indicated.NSC II cells exhibit distinct secretory cycles. Early in the cycle the cytoplasm becomes phloxinophilic and progresses to a distinct fuchsinophilic stage. Small homogeneous irregular inclusions are found in the axon hillock during the latter stages of the cycle. Histochemical tests suggest the presence of a carbohydrate and strong acidic groups which may have been derived from S-S or S-H groups. There are small cells present which appear to be immature neurosecretory cells.NSC III cells are characterized by a perinuclear ring of cytoplasm which is stained by chrome alum hematoxylin but not by paraldehyde fuchsin. A secretory cycle may also be present in this cell type.The three cell types presumed to be ordinary neurons exhibit no particular staining affinity for the stains or tests used in this study.This study was supported in part by a grant from the Central Fund for Research of the Pennsylvania State University.     

Comparison of the autogenous potentials of newly-emerged anautogenous female mosquitoes

The autogenous potentials in several species of newly emerged anautogenous female mosquitoes were examined by homoplastic transplantion of resting stage ovaries into newly-emerged females. Ovaries from 4-day-old female Culex pipiens quinquefasciatus, with follicles at stage Ib, were all activated in the newly emerged hosts and deposited some yolk in the oocyte. In addition, some of the implants (5-10%) matured. Transplanted C. tritaeniorhynchus ovaries also started to develop beyond the Ib resting stage, but never grew beyond the IIb stage. Resting Armigeres subalbatus and Aedes albopictus ovaries, both follicles having already reached the IIa-IIb stages, did not show any further growth after being implanted into newly emerged hosts. However, resting Ae. albopictus ovaries matured when heteroplastically transplanted into Culex pipiens molestus (74%) or into Culex pipiens pallens (5%). In addition, Ae. albopictus females, acting as hosts, could mature transplanted C. p. molestus (23%) and C. p. pallens ovaries (10%). Thus, the above specific differences in autogenous potentials seem to reside not only in the internal environment of the hosts, but also in the ovaries themselves. Development of the host ovaries in the above homoplastic and heteroplastic transplantations were usually suppressed by the implanted donor ovaries and failed to reach the resting stages of each species.     

Ultrastructure of the neurosecretory system of the Colorado potato beetle,Leptinotarsa decemlineata (Say)

Summary The ultrastructure of seven types of neurosecretory cells (NSC) in the medial and lateral groups of the protocerebrum is described. The differences among cell types established earlier by light microscopy parallel differences in size and appearance of the neurosecretory particles observed in electron micrographs. No relationship was found between the affinity for Gomori's paraldehyde fuchsin stain and the nature of the particles.The secretions of the A-, A₁-, and C-types of NSC of the medial group are characterized by electron-dense neurosecretory granules of 1250 Å dia., medium-dense granules of 2100 Å, and electron-lucent vesicles of 1700 Å, respectively. The L-type NSCof the lateral group contain smaller (1300 Å) or larger (1700 Å) neurosecretory granules. The medial B- and E-types of NSC and the lateral L_B-type contain granulated vesicles (1200 Å) of the same appearance. These cell types differ in other respects and most likely have separate functions.The author wishes to thank the Laboratory of Virology of the Agricultural University for the use of the electron microscope, Mr. J. Groenewegen and Miss J. van Rinsum for technical assistance, and Professor J. Lattin for correcting the English text. Part of the work has been done while the author was in the service of the Netherlands Organization for the Advancement of Pure Research (ZWO, grant 942-48), and the National Council for Agricultural Research (TNO).     

Neurosecretory activity during regeneration in the oligochaete annelid Tubifex tubifex

Following amputation of the posterior half of Tubifex, all the nerve cells stainable by paraldehyde fuchsin (NF) immediately discharged. Fifteen minutes later, some stained NFs reappeared. Their number increased, reached a peak far above the normal level, then slowly decreased. This peak occurred 8 hours after amputation in the brain, after 3-4 days in the ventral ganglia. A similar cycle took place during the anterior regeneration of Tubifex whose four anterior segments were previously cut away. Following a double amputation (behind the first or fourth segment and at half length), the stimulation of the neurosecretory activity was even clearer, the number of NFs reaching a higher and earlier peak. Correlatively, the regenerative histogenesis was accelerated in the caudal blastema.     

Isolation of the spirochaete Borrelia afzelii from the mosquito Aedes vexans in the Czech Republic

During the years 1993–1995, a total of 3580 culicine mosquitoes of six species were collected in South Moravia, Czech Republic, and examined by dark-field microscopy for the presence of borreliae. Females of Aedes cantans , Ae. sticticus , Ae. vexans , Culex pipiens and Cx pipiens biotype molestus (but not Ae. geniculatus or Culiseta annulata ) harboured spirochaetes, the frequencies ranging from 0.7% to 7.8%. One isolate (BR-53) from Ae. vexans was identified as Borrelia afzelii genospecies. The potential role of mosquitoes in the epidemiology of Lyme borreliosis should be investigated.     

中国尖音库蚊复合组生物分类学的研究(英文)

本文对我国尖音库蚊复合组(Culex pipiens complex)进行了研究,其中包括从新疆采集尖音库蚊。建立实验室株,进行杂交、不同地理株形态学数值分析、杂种F_1代形态学的数值分析,表皮碳氢化合物、体内脂肪酸和单糖气相色谱分析以及卵蛋白质的电泳分析等。在我国首次发现了骚扰库蚊。根据研究结果,我国异域分布的尖音库蚊复合组实际上包含着同一个种的4个亚种,即指名亚种(Culex pipiens pipiens),淡色库蚊亚种(Culex pipiens pallens)、致倦库蚊亚种(Culex pipiens quinquenfasciatus)和骚扰库蚊亚种(Culex pipiens molestus)。致倦库蚊不应作为独立的蚊种,淡色库坟也不应作为所谓“中间型”。在亲缘关系上根据形态学特征聚类分析等结果表明,尖音库蚊与致倦库蚊的距离较远。