Neurosecretory Research Articles

Parasite reliance on its host gut microbiota for nutrition and survival

Studying the microbial symbionts of eukaryotic hosts has revealed a range of interactions that benefit host biology. Most eukaryotes are also infected by parasites that adversely affect host biology for their own benefit. However, it is largely unclear whether the ability of parasites to develop in hosts also depends on host-associated symbionts, e.g., the gut microbiota. Here, we studied the parasitic wasp Leptopilina boulardi (Lb) and its host Drosophila melanogaster. Results showed that Lb successfully develops in conventional hosts (CN) with a gut microbiota but fails to develop in axenic hosts (AX) without a gut microbiota. We determined that developing Lb larvae consume fat body cells that store lipids. We also determined that much larger amounts of lipid accumulate in fat body cells of parasitized CN hosts than parasitized AX hosts. CN hosts parasitized by Lb exhibited large increases in the abundance of the bacterium Acetobacter pomorum in the gut, but did not affect the abundance of Lactobacillus fructivorans which is another common member of the host gut microbiota. However, AX hosts inoculated with A. pomorum and/or L. fructivorans did not rescue development of Lb. In contrast, AX larvae inoculated with A. pomorum plus other identified gut community members including a Bacillus sp. substantially rescued Lb development. Rescue was further associated with increased lipid accumulation in host fat body cells. Insulin-like peptides increased in brain neurosecretory cells of parasitized CN larvae. Lipid accumulation in the fat body of CN hosts was further associated with reduced Bmm lipase activity mediated by insulin/insulin-like growth factor signaling (IIS). Altogether, our results identify a previously unknown role for the gut microbiota in defining host permissiveness for a parasite. Our findings also identify a new paradigm for parasite manipulation of host metabolism that depends on insulin signaling and the gut microbiota.

Spatial Transcriptomics Reveal Potential Sex Differences in Gene Expression of the Supraoptic Nucleus

The supraoptic nucleus (SON) of the hypothalamus contains magnocellular neurosecretory cells that play a key role in the regulation of fluid and electrolyte homeostasis. Although there are many well‐known sexually dimorphic regions of the hypothalamus, little is known about whether this is also true for the SON and whether there are sex differences in SON gene expression. Our study aims to address this knowledge gap by leveraging spatially‐resolved transcriptomics to better visualize gene expression profiles of cells in the SON of male and female rats and gain insight on their physiological functions without sacrificing morphological context. Visium Spatial Gene Expression (10x Genomics) was used to obtain spatially‐resolved gene expression data for the SON of adult male (n=3) and female (n=3) Sprague‐Dawley rats. Briefly, each brain was sectioned at 10μm thickness to collect coronal sections (~4x4mm) containing the SON and other brain structures. Each section was then mounted on the capture areas of Visium slides containing probes that bind mRNA. Next, the sections underwent the workflow as follows: 1) sample staining and imaging, 2) cDNA library preparation, 3) sequencing, and 4) analysis/data visualization. Data were analyzed using 10x Genomics’ Space Ranger and Loupe Browser applications to overlay spatial gene expression data with reference images. Results show that gene cluster analysis successfully differentiated myelinated fiber tracts from nuclei and identified several distinct neuronal populations in the coronal brain sections from both male and female rats. From the list of significant genes after performing differential expression analysis on the SON region using 10x Genomics’ Loup Browser, 74/157 genes were unique to the males, 228/311 genes were unique to the females, and 83 genes (e.g., Avp and Oxt) were common to both sexes. Preliminary Gene Ontology (GO) Enrichment and pathway analyses revealed GO terms and pathways related to: 1) homeostatic processes, regulation of peptide hormone secretion, and ion transport for the common genes; 2) ribosomes and translation for the significant genes unique to males; and 3) neurotransmitter release cycle and synaptic transmission for the significant genes unique to females, as some examples. These spatially‐resolved transcriptomic data suggest potential sex differences in SON gene expression that may be associated with processes and pathways important for osmoregulation and fluid balance. Future spatial transcriptomic studies will investigate changes in SON gene expression that contribute to sex differences in cellular mechanisms involved in body fluid homeostasis and possibly pathophysiology.

The Neuronal Circuit of the Dorsal Circadian Clock Neurons in Drosophila melanogaster.

Drosophila’s dorsal clock neurons (DNs) consist of four clusters (DN1as, DN1ps, DN2s, and DN3s) that largely differ in size. While the DN1as and the DN2s encompass only two neurons, the DN1ps consist of ∼15 neurons, and the DN3s comprise ∼40 neurons per brain hemisphere. In comparison to the well-characterized lateral clock neurons (LNs), the neuroanatomy and function of the DNs are still not clear. Over the past decade, numerous studies have addressed their role in the fly’s circadian system, leading to several sometimes divergent results. Nonetheless, these studies agreed that the DNs are important to fine-tune activity under light and temperature cycles and play essential roles in linking the output from the LNs to downstream neurons that control sleep and metabolism. Here, we used the Flybow system, specific split-GAL4 lines, trans-Tango, and the recently published fly connectome (called hemibrain) to describe the morphology of the DNs in greater detail, including their synaptic connections to other clock and non-clock neurons. We show that some DN groups are largely heterogenous. While certain DNs are strongly connected with the LNs, others are mainly output neurons that signal to circuits downstream of the clock. Among the latter are mushroom body neurons, central complex neurons, tubercle bulb neurons, neurosecretory cells in the pars intercerebralis, and other still unidentified partners. This heterogeneity of the DNs may explain some of the conflicting results previously found about their functionality. Most importantly, we identify two putative novel communication centers of the clock network: one fiber bundle in the superior lateral protocerebrum running toward the anterior optic tubercle and one fiber hub in the posterior lateral protocerebrum. Both are invaded by several DNs and LNs and might play an instrumental role in the clock network.

NEUROSECRETION IN OLIGOCHAETES - A LESS STUDIED SUBJECT IN INVERTEBRATE RESEARCH

The concept of neurosecretion was first developed by Ernst Scharrer (1905-1965) and Berta Scharrer (1906-1995) between 1928 and 1937. The neurosecretory cells are neuronal in structure and glandular in function. Typically, the neurosecretory cells are “tear drop” in appearance with a long axon. The neurosecretory products (neurosecretion/neurohormone) are proteinaeous in nature which is consistent with the ultrastructure of the cells
 In earthworm species, irrespective of their ecological categories, the neurosecretory cells in their distribution, structure and functions in the central nervous system are more or less similar in different species of earthworms. The central nervous system in tropical earthworms reveals mainly two types of neurosecretory cells: A type (outer cortical tier of deeply stained AF positive cells with distinct axonal processes) and B type (light stained cells lying in between A type cells and central fibrous neuropile). In addition to A and B cells, an “islet” of neurosecretory cells called S cells or Hubl cells are present at the base of circum esophageal connectives of subesophageal ganglia. Both A and B cells discharge their secretion (neurohormone) at the margin of highly vascularised fibrous neuropile called “zone of accumulation” which may be considered as “elementary neuro-haemal organ”. Neurosecretion in Oligochaetes has great role in food intake, osmoregulation, thermal acclimation, regeneration, copulation and cocoon production.

PACAP/GCGa Is an Important Modulator of the Amphioxus CNS-Hatschek's Pit Axis, the Homolog of the Vertebrate Hypothalamic-Pituitary Axis in the Basal Chordates.

The Hatschek’s pit in the cephalochordate amphioxus, an invertebrate deuterostome basal to chordates is suggested to be the functional homolog structure of the vertebrate adenohypophysis based on anatomy and expression of homologous neuroendocrine genes. However, the endocrine potential of the cephalochordate Hatschek’s pit remains to be demonstrated as well as the physiological actions of the secreted neuropeptides. In this study, we have explored the distribution and characterize the potential function of the amphioxus PACAP/GCG precursor, which is the ortholog of the hypothalamic PACAP neuropeptide in vertebrates. In amphioxi, two PACAP/GCG transcripts PACAP/GCGa and PACAP/GCGbc that are alternative isoforms of a single gene with different peptide coding potentials were isolated. Immunofluorescence staining detected their expression around the nucleus of Rohde, supporting that this structure may be homologous of the neurosecretory cells of the vertebrate hypothalamus where abundant PACAP is found. PACAP/GCGa was also detected in the infundibulum‐like downgrowth approaching the Hatschek’s pit, indicating diffusion of PACAP/GCGa from the CNS to the pit via the infundibulum‐like downgrowth. Under a high salinity challenge, PACAP/GCGa was upregulated in amphioxi head and PACAP/GCGa treatment increased expression of GHl in Hatschek’s pit in a dose-dependent manner, suggesting that PACAP/GCGa may be involved in the regulation of GHl via hypothalamic-pituitary (HP)-like axis similar as in the vertebrates. Our results support that the amphioxus Hatschek’s pit is likely to be the functional homolog of pituitary gland in vertebrates.

Drosophila TRPγ is required in neuroendocrine cells for post-ingestive food selection.

The mechanism through which the brain senses the metabolic state, enabling an animal to regulate food consumption, and discriminate between nutritional and non-nutritional foods is a fundamental question. Flies choose the sweeter non-nutritive sugar, L-glucose, over the nutritive D-glucose if they are not starved. However, under starvation conditions, they switch their preference to D-glucose, and this occurs independent of peripheral taste neurons. Here, we found that eliminating the TRPγ channel impairs the ability of starved flies to choose D-glucose. This food selection depends on trpγ expression in neurosecretory cells in the brain that express diuretic hormone 44 (DH44). Loss of trpγ increases feeding, alters the physiology of the crop, which is the fly stomach equivalent, and decreases intracellular sugars and glycogen levels. Moreover, survival of starved trpγ flies is reduced. Expression of trpγ in DH44 neurons reverses these deficits. These results highlight roles for TRPγ in coordinating feeding with the metabolic state through expression in DH44 neuroendocrine cells.

A local insulin reservoir in Drosophila alpha cell homologs ensures developmental progression under nutrient shortage

Insulin/insulin-like growth factor (IGF) signaling (IIS) controls many aspects of development and physiology. In Drosophila, a conserved family of insulin-like peptides called Dilps is produced by brain neurosecretory cells, and it regulates organismal growth and developmental timing. To accomplish these systemic functions, the Dilps are secreted into the general circulation, and they signal to peripheral tissues in an endocrine fashion. Here, we describe the local uptake and storage of Dilps in the corpora cardiaca (CC), an endocrine organ composed of alpha cell homologs known to produce the glucagon-like adipokinetic hormone (AKH). We show that Dilp uptake by the CC relies on the expression of an IGF-binding protein called ImpL2. Following their uptake, immunogold staining demonstrates that Dilps are co-packaged with AKH in dense-core vesicles for secretion. In response to nutrient shortage, this specific Dilp reservoir is released and activates IIS in a paracrine manner in the prothoracic gland. This stimulates the production of the steroid hormone ecdysone and initiates entry into pupal development. We therefore uncover a sparing mechanism whereby insulin stores in CC serve to locally activate IIS and the production of ecdysone in the PG, accelerating developmental progression in adverse food conditions.

Calmidazolium induces a decrease in nicotine-induced currents and intracellular calcium levels after pulse application of nicotine onto insect neurosecretory cells

Dorsal unpaired median (DUM) neurons, are a class of insect neurosecretory cells, which are involved in the control of several functions, such as excretion and reproduction, or the release of neurohormones. Previous studies demonstrated that they express different nicotinic acetylcholine receptor subtypes, in particular α-bungarotoxin-insensitive receptors, with nAChR1 and nAChR2 subtypes. Here, we demonstrated that pulse application of 1 mM nicotine (300 ms pulse duration) induced inward currents which were reduced under bath application of 15 µM calmidazolium, a calmodulin inhibitor. Bath application of 0.5 µM α-bungarotoxin had no effect on calmidazolium action, suggesting that it could have an indirect effect through α-bungarotoxin-insensitive receptors. Indeed, nicotine-evoked currents were reduced by 10 µM d-tubocurarine, and completely blocked by 5 µM mecamylamine, which affected nAChR1 and nAChR2 subtypes, respectively. Our results demonstrated that nAChR2 subtypes are involved in the indirect effect of calmidazolium. Moreover, we found that this calmidazolium effect was associated to a strong reduction in intracellular calcium levels after pulse application of 1 mM nicotine. Thus, compared to previous studies on mammalian cells, calmidazolium did not cause an increase in intracellular calcium levels in DUM neurons, suggesting that different calcium mechanisms are involved in the calmidazolium effect.

Lactate-Dependent Cross-Talk Between Astrocyte and GnRH-I Neurons in Hypothalamus of Aged Brain: Decreased GnRH-I Transcription.

GnRH-I produced by hypothalamic neurosecretory cells is considered a master regulator of mammalian reproduction. Although GnRH-I transcription is well studied, the effect of ageing on transcriptional regulation of GnRH-I has not yet been explored. Here, we elucidate the effects of ageing on the metabolic environment like lactate level and TNF-α and how these affect GnRH-I transcription. Using pathway analysis of transcriptomic data, we found that lactate is upregulated in ageing astrocytes due to the downregulation of cellular respiration pathways possibly resulting in greater pyruvate concentration for lactate production. This lactate could then be shuttled into neurons where it would affect GnRH-I transcription. We showed that supra-physiological level of lactate in young mouse brain can mimic metabolic disturbances in the old brain and cause downregulation in GnRH-I transcription at a young age. In particular, we found upregulation of GnRH-I repressors in the young brain treated with high levels of lactate similar to old brain. Hence, this confirmed that aged metabolic environment can affect GnRH-I transcription even in the young brain. Further downstream analysis using the TRUST database showed NF-Kb signalling which lies downstream of both lactate and TNF-α as being capable of upregulating GnRH-I repressors. Since NF-Kb signalling has been shown in our study as well as others to be induced by TNF-α during ageing, it is likely that GnRH-I transcriptional regulation is mediated through these pathways. Thus, we formed a model for explaining the downregulation of GnRH-I transcription during ageing through differential expression of its TFs in an aged metabolic environment.

Histological, histochemical, ultrastructural and immunohistochemical identification and characterization of the neurosecretory cells of the adult rabbit's adrenal medulla.

The present investigation was conducted on the adrenal glands of 40 adult New Zealand rabbits of both sexes to characterize and identify the histological, histochemical, ultrastructural, and immunohistochemical features of the neurosecretory cells of the adrenal medulla. The obtained specimens of adrenal medulla were subjected to routine histological techniques and then stained with different histological stains, including general, non-specific, specific, and highly specific stains for neurosecretory cells, in addition to immunohistochemical reactions. The obtained results showed two types of adrenal medullary neurosecretory cells containing secretory granules (SGs) of different electron densities: adrenaline and noradrenaline (NA) secreting cells. These secretory granules showed a strong positive reaction to the Grimelius silver impregnation technique. Sections stained with Gomori's chrome alum haematoxylin stain, and the secretory granules showed a strong dark blackish-blue positive colour. The medullary cells showed typical chromaffin reactions when stained by H&E and Giemsa stains after formol dichromate 'Ortha's fluid' fixation. The noradrenaline secretory granules gave a strong positive Schmorl's test, while the adrenaline ones showed a moderate reaction. Immunohistochemically, the adrenal medullary cells were subjected to anti-chromogranin A (CHGA) antibody using the PAP technique, which gave positive reactions.

Exploring the role of glycoprotein hormone GPA2/GPB5 in the medically important insect, Rhodnius prolixus

Glycoprotein hormones are formed by the heterodimerization of alpha and beta subunits. In vertebrates, there are five glycoprotein hormones, four of which have a common alpha subunit (GPA1) bound to a specific beta subunit (GPB1, GPB2, GPB3, or GPB4), and the fifth, thyrostimulin, is formed by the dimerization of GPA2 and GPB5 subunits. These hormones mediate physiological events such as development, metabolism, and reproduction, although the functional role of thyrostimulin in vertebrates has not been fully elucidated. Recent reports in invertebrates, specifically in holometabolous insects, suggest that GPA2/GPB5 plays a critical role in development, diuresis, and reproduction. In this study, we clone and characterize the transcripts for the glycoprotein hormone GPA2/GPB5 and its receptor (LGR1) in fifth instar Rhodnius prolixus, a hemimetabolous insect vector of Chagas disease. Sequence analyses reveals considerable identity and similarity between GPA2/GPB5 and LGR1 and those reported in other arthropod species. Quantitative PCR (qPCR) shows that both subunit transcripts, GPA2 and GPB5, and LGR1 transcripts are present in a variety of tissues, with greatest expression of the subunits in the central nervous system (CNS) and highest LGR1 expression in the Malpighian tubules (MT). Results from temporal qPCR analyses reveal a decrease in transcript expression 24 h after feeding, followed by an increase as the days post-feeding advance. Using immunohistochemistry, we show that GPB5 is expressed throughout the CNS, and importantly is present in neurosecretory cells in the brain and abdominal neuromeres and their neurohemal organs, indicating a neurohormonal role for this signaling pathway. A reduction in LGR1 transcript expression (via RNA interference) led to a greater weight loss and mortality rate in unfed insects. In addition, when a blood meal is offered, the insects with reduced LGR1 consume a significantly smaller blood meal and have higher mortality rates as the days post-feeding advance. Overall, the results suggest that the GPA2/GPB5 signaling pathway may play roles during a prolonged unfed state and in feeding-related events.

The influence of melatonin on stress-induced ultramicroscopic changes of the neurons of the supraoptic nuclei of the rat hypothalamus

The aim: to study the effect of melatonin on the ultrastructural state of the supraoptic nuclei of the hypothalamus of rats under immobilization stress.Materials and methods. The experiments were performed on non-linear male white rats weighing 200-220 g. The animals were divided into 3 study series, in each of which the biomaterial was collected at 2 p.m. and at 2 a.m. using electron microscopic method. Long immobilization stress was simulated by keeping rats in special plastic penal cages for 6 hours daily for 7 consecutive days. Melatonin (Sigma, USA, 99.5% purification degree) at a dose of 0.5 mg/kg, in 1.0 ml of solvent (0.9% ethanol solution on physiologic saline) was injected daily, intraperitoneally.Results. When the animals were kept under the standard light regime, the ultrastructural organization of the hypothalamic nuclei at 2 p.m. indicated their low functional activity in comparison with the studies carried out at 2 a.m. Prolonged exposure of rats to immobilization stress was reflected in a significant rearrangement of the ultrastructural organization of supraoptic nuclei of the hypothalamus. The established changes can be considered as a manifestation of neurosecretory activity suppression, a decrease in neurosecretase production by hypothalamic neurons. Melatonin injections against the background of immobilization stress resulted in relative normalization of ultrastructural state of neurons of supraoptic nuclei of the hypothalamus of animals. In particular, studies at 2 a.m. revealed light neurosecretory cells containing a large nucleus, it was pyknotically altered. Karyolema invaginations, euchromatin dominance in the nucleus were observed. Heterogeneous changes were observed on the part of mitochondria. Enlarged tubules of granular endoplasmic reticulum were seen. At the same time, a small number of ribosomes and few hormonal granules were noticeable in neuroplasm. The mentioned picture of neurosecretory cells reflects a relative improvement in their electron microscopic state, which is evidenced by the appearance of neurosecretory granules. However, the ultrastructure of other organelles of the studied neurons indicates a depleted state caused by prolonged immobilization.Conclusions. 1. In animals under standard photoperiod conditions, the structural organization of supraoptic neurons of the hypothalamic nuclei during the nighttime of the experiment reflects the intensity of intracellular synthesizing processes (at 2 a.m.). A decrease in the activity of the structures under study is noted during the daytime. 2. Under immobilization stress, the ultrastructural organization of the above neurons indicates a pronounced disturbance of reactive nature with the signs of decreased functional ability of the structures and the phenomena of edema and destruction during the period of observation. 3. Melatonin injections against the background of immobilization stress led to a relative improvement in the ultrastructural state of the animals’ hypothalamic nuclei neurons, which is evidenced by the appearance of neurosecretory granules. However, the ultrastructure of other organelles of the studied neurons indicated a depleted state caused by prolonged immobilization.

Transgenic approaches to opening up new fields of vasopressin and oxytocin research.

Transgenic approaches have been applied to generate transgenic rats that express exogenous genes in arginine vasopressin (AVP)- and oxytocin (OXT)-producing magnocellular neurosecretory cells (MNCs) of the hypothalamic-neurohypophyseal system (HNS). First, the fusion gene that expresses AVP-enhanced green fluorescent protein (eGFP) and OXT-monomeric red fluorescent protein 1 (mRFP1) was used to visualize AVP- and OXT-producing MNCs and their axon terminals in the HNS under fluorescence microscopy. Second, the fusion gene that expresses c-fos-eGFP and c-fos-mRFP1 was used to identify activated neurons physiologically in the central nervous system, including MNCs, circumventricular organs and spinal cord. In addition, AVP-eGFP x c-fos-mRFP1 and OXT-mRFP1×c-fos-eGFP double transgenic rats were generated to identify activated AVP- and OXT-producing MNCs using appropriate physiological stimuli. Third, the fusion gene that expresses AVP-chanelrhodopsin 2 (ChR2)-eGFP and AVP-hM3Dq-mCherry was used to activate AVP- and OXT-producing MNCs by optogenetic and chemogenetic approaches. In each step, these transgenic approaches in rats have provided new insights on the physiological roles of AVP and OXT not only in the HNS, but also in the whole body. In this review, we summarize the transgenic rats that we generated, as well as related physiological findings.

Mechanism and function of phasic firing in vasopressin-releasing magnocellular neurosecretory cells.

Magnocellular neurosecretory cells that release vasopressin (MNCVP ) from axon terminals in the neurohypophysis display a unique pattern of action potential firing termed phasic firing. Under basal conditions, only a small proportion of MNCVP display spontaneous phasic firing. However, acute and chronic conditions that stimulate vasopressin release, such as hemorrhage and dehydration, greatly enhance the number of MNCVP that fire phasically. Phasic firing optimizes VP neurosecretion at axon terminals by allowing action potential broadening to promote calcium-dependent frequency-facilitation, at the same time as preventing the secretory fatigue caused by spike inactivation that occurs during prolonged continuous stimulation. This review provides an update on our mechanistic understanding of these processes and highlights important gaps in our knowledge that must be addressed in future experiments.

Study of triclabendazole effects on Fasciola hepatica’s life-supporting organs, spines and suckers responsible for stable position of the parasite in the host

The purpose of the research is to study triclabendazole effects on the Fasciola’s life-supporting organs, spines and suckers which are responsible for stable position of the parasite in the host.Materials and methods. The study material was trematodes Fasciola hepatica (Linneus 1758, family Fasciolidae Railliet 1895), which were collected after the action of triclabendazole (fasinex) (chemically 5-chloro-6-(2,3-dichlorophenoxy)-2-methylthiobenzimidazole)on the 7th day after the drug administered at a single dose of 10 mg/kg for the Active Substance in the treatment of ovine fasciolosis. F. hepatica from untreated animals served as control. Mature F. hepatica collected after treatment with triclabendazole, and marita from the control groups were dehydrated in ascending alcohol series for 1–2 days after fixation; then passed through a mixture of chloroform and absolute alcohol (in a ratio of 1:1), and through pure chloroform in two portions for 10–15 minutes. The material was then soaked in a mushy mixture of chloroform and paraffin in a thermostat at 37 °C for 12–18 hours, and in paraffin in a thermostat at 56 °C for 30–45 minutes; and then embedded in paraffin with added wax. The resulting paraffin blocks were broken down into serial sections of 5–7 μm thick, then stained and examined under a light microscope.Results and discussion. Pathomicromorphological analysis of F. hepatica’s spines and suckers, organs that come into adhesive contact with the host organism revealed destructive changes in them after the action of triclabendazole. After the action of triclabendazole on fascioles, the spines look enlarged and swollen, and have a more rounded shape and some changes in color, absorbing eosin in greater concentration. The muscle fibers of the fascioles’ oral and abdominal suckers also look swollen after the action of triclabendazole. Although the musculature of the F. hepatica’s pharynx retained its structure, it has changes. It thickened sharply, which is clearly visible on the transverse and longitudinal sections of the helminths; neurosecretory cells are destroyed, and voids are observed in their place.

A nutrient-responsive hormonal circuit mediates an inter-tissue program regulating metabolic homeostasis in adult Drosophila

Animals maintain metabolic homeostasis by modulating the activity of specialized organs that adjust internal metabolism to external conditions. However, the hormonal signals coordinating these functions are incompletely characterized. Here we show that six neurosecretory cells in the Drosophila central nervous system respond to circulating nutrient levels by releasing Capa hormones, homologs of mammalian neuromedin U, which activate the Capa receptor (CapaR) in peripheral tissues to control energy homeostasis. Loss of Capa/CapaR signaling causes intestinal hypomotility and impaired nutrient absorption, which gradually deplete internal nutrient stores and reduce organismal lifespan. Conversely, increased Capa/CapaR activity increases fluid and waste excretion. Furthermore, Capa/CapaR inhibits the release of glucagon-like adipokinetic hormone from the corpora cardiaca, which restricts energy mobilization from adipose tissue to avoid harmful hyperglycemia. Our results suggest that the Capa/CapaR circuit occupies a central node in a homeostatic program that facilitates the digestion and absorption of nutrients and regulates systemic energy balance.

Progress in the characterization of insulin-like peptides in aphids: Immunohistochemical mapping of ILP4

Aphids were the first animals described as photoperiodic due to their seasonal switch from viviparous parthenogenesis to sexual reproduction (cyclical parthenogenesis) caused by the shortening of the photoperiod in autumn. This switch produces a single sexual generation of oviparous females and males that mate and lay diapausing cold-resistant eggs that can overcome the unfavourable environmental conditions typical of winter in temperate regions. Previous studies have hinted at a possible implication of two insulin-like peptides (ILP1 and ILP4) in the aphid seasonal response, changing their expression levels between different photoperiodic conditions. Moreover, in situ localization of their transcripts in particular neurosecretory cells (NSCs) in the aphid brain supported the idea that these neuropeptides could correspond to the formerly called virginoparin, an uncharacterized factor originally proposed to be transported directly to the aphid embryos to promote their development as parthenogenetic individuals. To further investigate the fate of these ILPs, we raised a specific antiserum against one of them (ILP4) and mapped this neuropeptide by immunohistochemistry (IHC) in Acyrthosiphon pisum and Megoura viciae aphids. Coincident with in situ localization, our results show that ILP4 is synthesized in two groups (one in each brain hemisphere) of four neurosecretory cells in the pars intercerebralis (NSC group I) and then it is transported outside the brain to the corpora cardiaca. From there, three nerves (two laterals and one medial) transport it to the abdomen. Although no precise site of release has been found, the terminations of these nerves near the germaria would be compatible with the proposal of a direct connection between group I of NSCs and the reproductive system by localized release. In addition, we detected some collateral arborizations originating from the eight NSCs going to the pars lateralis, where clock neurons and some photoreceptors have been previously localized, suggesting a possible communication between the circadian and photoperiodic systems.

The adverse effects of injected functionalized multi-walled carbon nanotube (f-MWCNT) on in vivo neurosecretory brain cells of Jamaican field cricket, Gryllus assimilis.

Carbon nanotubes (CNTs) have been increasingly more prevalent due to their use in product technology owing to their exceptional electrical and thermal conductivity and tensile strength because of their nanostructure and strength of the bonds among carbon atoms. The potential increase of CNTs in the environment is a concern, and studies to assess the toxic effects of these nanomaterials (NMs) are needed. However, so far, most of the studies are focused on aquatic species and much less is understood about the effects of NM in terrestrial organisms. This investigation used a functionalized multi-walled carbon nanotube (f-MWCNT) and the Jamaican cricket Gryllus assimilis to assess the effects of this NM. Cricket nymphs were injected with f-MWCNT suspension-at three different concentrations. The insecticide Fipronil was used as a positive control. Survival was monitored, and histological analysis was made in the brains. Pyknotic cells were quantified in two brain regions, a neurosecretory called Pars intercerebralis (PI), and an associative region called mushroom body (MB). No mortality was observed in any f-MWCNT concentration tested. A significant increase in pyknotic cells was observed as sub-lethal effect for the intermediate concentration of f-MWCNT, at PI, while any significant change was observed at the Kenyon cells of the MB. These results are discussed in the context of agglomeration and dispersion of the f-MWCNT at different concentrations, and availability of the f-MWCNT on the circulatory system, as well as the natural decay of pyknotic cells with time and different patterns of adult cricket neurogenesis. Our results showed that f-MWCNT had negative effects in the neurosecretory region of the brain.

Neuroendocrine control of cocoon production in native earthworm Perionyx ceylanensis subjected to seasonal variation

Aim: The aim of the present study was to observe the role of cerebral ganglionic neurosecretory cells (NSCs) during cocoon production in native earthworm species Perionyx ceylanensis subjected to amputation and seasonal variations. Methodology: Histological studies (using Aldehyde Fuchsin and Chrome Alum Haematoxylin Phloxin stain) were carried out on brain NSCs in the two groups of earthworms (Group I and Group II) maintained in earthen culture pots (2L) with cowdung as food. Group I comprised of brain amputed earthworms was subjected to observe the role of brain NSCs in production of cocoon and Group II earthworms subjected to seasonal changes in the cerebral NSCs during cocoon production. Five replications were kept for Group I (1 individual per pot) and Group II (1 pair per pot). Results: Group I debrained earthworms started to lay cocoons from the 31st day following regeneration of cerebral ganglionic type A NSCs. In group II worms the highest neurosecretory activity was registered in the cerebral type A cells, especially during monsoon coinciding with the hike of cocoon generation. Interpretation: Appearance of type A NSCs in regenerated brain and peak of type A neurosecretory cell activity during peak reproductive period of earthworm species (as indicated by peak of cocoon production) indicates the possible role of cerebral type A NSCs in cocoon laying.

Photoperiodic control of electrophysiological properties of the caudo-dorsal cells in the pond snail, Lymnaea stagnalis.

Egg laying in the pond snail, Lymnaea stagnalis is regulated by the photoperiod; long-day conditions (16L8D) promote egg laying whereas medium-day conditions (12L12D) suppress it. In this snail, a caudo-dorsal cell hormone (CDCH) is produced by neurosecretory cells, CDCs in the cerebral ganglion (CG), and its release triggers ovulation and subsequent egg laying. However, the physiological basis for photoperiod-dependent egg laying remains unraveled. Here, we compared electrophysiological properties of CDCs between 16L8D and 12L12D using intracellular recording, and found that CDC excitability is higher in 16L8D than in 12L12D. Striking differences are as follows: (1) a shallower resting membrane potential in 16L8D than in 12L12D, and (2) a smaller threshold voltage (minimum depolarization from rest to elicit action potentials) in 16L8D than in 12L12D. Switching of the excitability can be a physiological basis of a photoperiod-dependent CDCH release. Simultaneous intracellular dye injection identified two morphological subtypes of CDCs, validating a previous report. Both types bear short lateral extensions in CG, some of which probably function as integration sites of photoperiodic inputs. In addition, we found two novel CDCH-immunoreactive cell groups (CDCCOM and SCm ) in the CG besides conventional CDCs and small cells expressing CDCH. The CDCCOM with cell bodies and fibers in the neurohemal commissure may be involved in triggering ovulation. Notably, the total number of CDCs is larger than that previously reported, the right CDC cluster with more cells than the left. Our findings are instructive in following the neurophysiology of photoperiodism in L. stagnalis.