Short-term captivity drives hypothalamic plasticity and asymmetry in wild-caught northern red bellied dace (Chrosomus eos).

Teleost fish are neuroplastic and are known to alter their brain morphology and behaviour in response to environmental change such as an increase in predation pressure. The hypothalamus plays a key role in regulating behavioural responses to predation risk. In this study, wild-caught northern red bellied dace (Chrosomus eos) developed smaller and less symmetric hypothalami when held in captivity for 14 days; both measures correlated with boldness in a latency to emerge test. This study's results highlight the potential impact of short-term holding conditions on brains and behaviour.

Serum biochemistry, liver histology and transcriptome profiling of bighead carp Aristichthys nobilis following different dietary protein levels.

Dietary protein plays a major role in determining the rate of fish growth and overall health. Given that the liver is an important organ for metabolism and detoxification, we hypothesized that optimal dietary protein levels may benefit liver function. Herein, we investigated the effects of dietary protein level on serum biochemistry, liver histology and transcriptome profiling of juvenile bighead carp Aristichthys nobilis fed for 8 weeks on a diet supplemented with high protein (HP, 40%), low protein (LP, 24%) or optimal protein (OP, 32%; controls). The results revealed a significant change in liver morphology in LP and HP groups compared with the OP group, coupled with increased serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activity. RNA sequencing (RNA-Seq) analysis of the liver transcriptome yielded 47 million high-quality reads using an Illumina platform, which were de novo assembled into 80,777 unique transcript fragments (unigenes) with an average length of 1021 bp. Subsequent bioinformatics analysis identified 878 and 733 differentially expressed unigenes (DEGs) in liver in response to LP and HP diets, respectively. KEGG enrichment analysis of DEGs identified immune and metabolism-related pathways, including Toll-like receptor signaling, PI3K-Akt signaling, NF-κB signaling, complement and coagulation, peroxisome, nitrogen metabolism, PPAR signaling, and glycolysis and gluconeogenesis pathways. Transcriptome profiling results were validated by quantitative real-time PCR for 16 selected DEGs. The findings expand our understanding of the molecular mechanisms underlying the effects of dietary protein level on liver function in bighead carp.

Detoxifying response in juvenile tench fed by selenium diet.

The effects of a selenium (Se) diet (1.0 mg Se kg⁻¹) were investigated on growth, accumulation and antioxidant response in juvenile Tinca tinca at three endpoints (0, 4 and 8 weeks). Growth and condition factor (K>1.5) for both control (0.25 mg Se kg⁻¹) and Se tench were not significantly affected. Se exposed fish exhibited the highest Se level in the kidney and the liver after 4 weeks. By feeding more Se the accumulation capacity of tench did not increase and a plateau, mainly for the liver, was thus reached. Se level remained almost constant in the muscle if compared to own control and for each endpoint. Superoxide dismutase activity in both tissues was not affected by Se supplementation and the higher catalase level in the kidney might support the hypothesis that the enzyme was adequate to remove the hydrogen peroxide production following Se exposure. However, supplemented diet with higher Se level could be critical for tench, as it may cause a lowering of glutathione peroxidase and glutathione reductase activities facilitating the onset of oxidative damage. The enhancement of thiol level and glutathione S-transferase activity, mainly in the liver, could be the signals of the only protection against the oxidative damage induced by Se.

Chelating efficacy of CaNa(2) EDTA on nickel-induced toxicity in Cirrhinus mrigala (Ham.) through its effects on glutathione peroxidase, reduced glutathione and lipid peroxidation.

In this age of modern biology, aquatic toxicological research has provided potential tools for ecotoxicologic investigations. Heavy metals primarily affect protein structures and induce a stress in the organisms. The present investigation was carried out to assess the effect of nickel chloride on the selected organs of the freshwater fish Cirrhinus mrigala and how CaNa(2) EDTA counters its effects as an antidote. Toxicity experiments were conducted for different exposure periods and also in certain tissues namely gill, liver, kidney and muscle. The total protein content, reduced glutathione, glutathione peroxidase and lipid peroxidation were found to be decreased in the nickel chloride treated tissues and the treatment with CaNa(2) EDTA+nickel chloride returned to near normal levels. Histopathological observations also revealed that after the administration of nickel chloride+CaNa(2) EDTA the chelator induced reduction in nickel toxicity. It has also contributed towards reduction in the pathological damage, thus enabling the organs to attain their near normal histological appearance. The present study shown that CaNa(2) EDTA is an effective chelating agent for the removal of nickel and it has proved efficient in restoring both the biochemical variables and pathological features immediately after a sub lethal exposure of nickel chloride in fish.

Health effects in fish of long-term exposure to effluents from wastewater treatment works.

Concern has been raised in recent years that exposure to wastewater treatment effluents containing estrogenic chemicals can disrupt the endocrine functioning of riverine fish and cause permanent alterations in the structure and function of the reproductive system. Reproductive disorders may not necessarily arise as a result of estrogenic effects alone, and there is a need for a better understanding of the relative importance of endocrine disruption in relation to other forms of toxicity. Here, the integrated health effects of long-term effluent exposure are reported (reproductive, endocrine, immune, genotoxic, nephrotoxic) . Early life-stage roach, Rutilus rutilus, were exposed for 300 days to treated wastewater effluent at concentrations of 0, 15.2, 34.8, and 78.7% (with dechlorinated tap water as diluent). Concentrations of treated effluents that induced feminization of male roach, measured as vitellogenin induction and histological alteration to gonads, also caused statistically significant alterations in kidney development (tubule diameter), modulated immune function (differential cell count, total number of thrombocytes), and caused genotoxic damage (micronucleus induction and single-strand breaks in gill and blood cells). Genotoxic and immunotoxic effects occurred at concentrations of wastewater effluent lower than those required to induce recognizable changes in the structure and function of the reproductive endocrine system. These findings emphasize the need for multiple biological end points in tests that assess the potential health effects of wastewater effluents. They also suggest that for some effluents, genotoxic and immune end points may be more sensitive than estrogenic (endocrine-mediated) end points as indicators of exposure in fish.

Cerebrospinal fluid-contacting neurons, sensory pinealocytes and Landolt's clubs of the retina as revealed by means of an electron-microscopic immunoreaction against opsin.

Opsin-immunoreactive sites of hypothalamic cerebrospinal fluid (CSF)-contacting neurons, pinealocytes and retinal cells were studied in various vertebrates (Carassius auratus, Phoxinus phoxinus, Triturus cristatus, Bombina bombina, Rana esculenta) by means of postembedding immuno-electron microscopy with the use of the protein A-gold labeling method. The retina of the rat served as a general reference tissue for the quality of the immunocytochemical reaction. A strong opsin immunoreaction (rat-antibovine opsin serum) was obtained in the rod-type outer segments of photoreceptors in the retina of all species studied. Cone-type outer segments exhibited only very few antigenic binding sites. In the pineal organ of the goldfish and the frog, outer segments of the photoreceptor cells displayed strong immunoreactivity. No immunoreaction was found in hypothalamic CSF-contacting neurons and Landolt's clubs of nerve cells of the bipolar layer of the retina. The morphological similarity between the ciliated dendritic terminal of the Landolt's club and the intraventricular dendritic ending of the CSF-contacting neurons is emphasized.

Forebrain connections in the goldfish support telencephalic homologies with land vertebrates.

Horseradish peroxidase injections into dorsomedial and dorsolateral regions of the goldfish (Carassius auratus) telencephalon demonstrate, by retrograde cell labeling, that the teleost telencephalon receives a pattern of projections from the thalamus remarkably similar to those of land vertebrates. The evidence provides support for a homology between the dorsomedial region and the corpus striatum of land vertebrates and a homology between two dorsolateral regions and the dorsal and medial pallium of land vertebrates.

Hypophysiotropic neurons in the goldfish hypothalamus demonstrated by retrograde transport of horseradish peroxidase.

The horseradish-peroxidase (HRP) technique was used to visualize the cell bodies of axons projecting to the goldfish pituitary. Following intravenous injections of HRP, HRP reaction products were observed in axons of the rostral pars distalis, proximal pars distalis, neurointermediate lobe, pituitary stalk and in axons coursing from the pituitary into the hypothalamus. HRP-labelled cells in the brain were localized in two regions only - the nucleus preopticus (NPO) pars magnocellularis and pars parvocellularis, and the nucleus lateralis tuberis (NLT) of the hypothalamus. These observations suggest that the NPO and NLT are the source of the neurosecretory innervation of the goldfish pituitary.

Cytological evidence for activation of neuroendocrine cells in the parvocellular preoptic nucleus of the goldfish hypothalamus following pharmacological adrenalectomy.

The nucleus preopticus (NPO) of the goldfish hypothalamus is composed of parvocellular (NPOpc) and magnocellular (NPOmc) neurosecretory neutrons. The cytology of NPOpc and NPOmc neurons was examined with light and electron microscopy following pharmacological adrenalectomy with the adrenocortical inhibitor, metopirone. After five days of metopirone administration, light microscopy revealed a significant increase in nuclear area of NPOpc, but not of NPOmc, neurons. Ultrastructural examination of NPOpc neurons revealed two cell types, PC1 and PC2 neurons, which could be distinguished by the relative abundance and the size of the neurosecretory granules in the cytoplasm. The ultrastructural appearance of the NPOmc neurons revealed a single cell type containing abundant neurosecretory granules. Following five days of metopirone administration, the ultrastructural appearance of the PC1 neurons indicated a state of enhanced secretory activity. Metopirone had no observable effect on the appearance of the PC2 or NPOmc neurons. These observations demonstrate that PC1 neurons are activated under the conditions of pharmacological adrenalectomy and suggest that the secretory activity of these neurons is inhibited by adrenocorticosteroids.

Specific effect of calcium ions on the calcium-sensitive cells of the pars intermedia in the goldfish.

Cytological changes in the calcium-sensitive (Ca-s) cells (formerly termed PAS-positive cells) of the pars intermedia were investigated in the goldfish after adaptation to deionized water (DW), with or without addition of sodium, potassium and magnesium. These ions were added as chloride salts at concentrations similar to those present in fresh water (FW). The marked stimulation of the Ca-s cells is not inhibited in DW supplemented with Na+ (0.35 mM/l), K+ (0.05 mM/l), and Mg2+ (0.2 mM/l) for a period of 24 days. The inhibition of the response to DW with calcium chloride (2 mM/l). These data show that chloride ions are not responsible for the regression of the Ca-s cells observed in goldfish kept in DW supplemented with calcium chloride. The effect of calcium ions on the Ca-s cells appears to be specific. These results support the hypothesis that the Ca-s cells synthesize a factor (hypercalcin?) involved in calcium regulation, and that its release is influenced by the calcium content of the environment. The role of the pars intermedia in calcium metabolism is strengthened by the present results. Biochemical data suggest the presence of a hypercalcemic factor in the pituitary of fish (Parsons et al. 1978) and are in agreement with the present cytological findings.

Hypothalamic lesions stimulating growth hormone cell activity in the goldfish.

The cytology of the growth-hormone (GH) cells of the goldfish pituitary were examined following electrothermic lesions of the anterior preoptic hypothalamus and telencephalon. Following lesions of the nucleus preopticus (NPO) light microscopy of the pituitary revealed a significant increase in the nuclear diameter and a degranulation of the GH cells. Lesions of the telencephalon anterior or dorsal to the NPO had no cytological effect on the GH cells. The ultrastructural appearance of the GH cells of NPO-lesioned fish was characterized by a marked degranulation of the cytoplasm and a proliferation of the rough endoplasmic reticulum indicative of enhanced secretory activity. The GH cells of the proximal pars distalis (PPD) are directly innervated by peptidergic (type A) and aminergic-like (type B) neurosecretory axons. Following lesions of the NPO, there was a marked reduction in the number of type A fibers in the PPD. These results suggest that the type A fibers innervating the GH cells originate in the NPO and act to inhibit the secretory activity of the GH cells.

PAS-positive cells of the pars intermedia are calcium-sensitive in the goldfish maintained in a hyposmotic milieu.

Cytological changes in the pars intermedia of the goldfish were investigated after adding calcium to deionized water (DW). In fish maintained in DW, the PAS-positive cells are highly stimulated in comparison to cells of fish kept in fresh water (FW). In DW supplemented with calcium at the same concentration as in FW (2 mM/l), the hyperactivity of the PAS-positive cells is prevented. When calcium ions are added 60 h before the animals are sacrificed, the PAS positive cells start to show signs of regression and their granules are stored: the release of the granular material appears to be suppressed by calcium. In the goldfish, the PAS-positive cells, homologous to a similar cell type in the eel, react only very weakly with the PAS technique. The name "calcium-sensitive cells" appears to be more appropriate in the goldfish for this particular cell type, secreting an unknown factor. This factor, different from the prolactin produced in the rostral pars distalis of the hypophysis, might be an equivalent of a "hypercalcin".

Morpho-histology of the hypothalamo-neurohypophyseal system in relation to gonadal maturation in Cirrhinus mrigala (Ham.), a freshwater Indian carp.

The hypothalamic neurosecretory centres, their pathway and the role of neurosecretory material responsible for gonadal maturation during its annual cycle is being reported here for the first time in an Indian freshwater major carp, Cirrhinus mrigala (Ham.).

Brain lesions and short-term endocrine effects of monosodium L-glutamate in goldfish, Carassius auratus.

Monosodium L-glutamate (MSG) was injected intraperitoneally into goldfish at a dosage of 2.5 mg/g body weight. At 24 h post-injection there was a marked hypertrophy and edema in the region of the nucleus lateralis tuberis (NLT) from the anterior margin of the pituitary stalk through to the posterior end of the NLT, irrespective of the sex of the goldfish. A similar hypertrophy and edema occurred ventral to the anterior commissure in the preoptic region in the anterior-ventral nucleus preopticus periventricularis (NPP). At 6 h post-injection a slight vacuolization was evident in these two regions, and at two days the hypertrophy and edema had abated from the extent observed at 24 h post-injection. At five and eight days post-injection only necrotic cells were found in the affected NLT region, but only a small band of necrotic cells was evident in the anterior-ventral preoptic region. No other brain lesions were evident. Serum levels of gonadotropin (GtH) were increased at 6 h, 24 h, and two days after treatment with MSG, but were similar to control values at five, seven and eight days after MSG in male and female goldfish. Exocytosis of small dark secretory granules in gonadotrophs was evident at 24 h after MSG in a fish with a somewhat greater increase in serum GtH than usually found. The time course of increased serum GtH levels postinjection of MSG is consistent with the observed time course of hypertrophy and atrophy of NLT neurons; the increase in serum levels of GtH is interpreted to reflect a stimulation of release of GtH-releasing factor from neurons in the NLT. Electron microscope investigation indicates that prolactin cells have increased secretory and synthetic activity from 24 h through to seven days post-injection of MSG. The mechanism for stimulation of the prolactin cells by MSG is not known. No other changes in activity of adenohypophysial secretory cells were found.

Immunocytochemical identification of enkephalinergic neurons in the hypothalamic magnocellular preoptic nucleus of the goldfish, Carassius auratus.

Our immunocytochemical investigation of the magnocellular neuroendocrine system in the goldfish hypothalamus reveals enkephalin (ENK)-containing neurons interspersed among the vasotocin (VT)- and isotocin (IT)-containing neurons of the preoptic nucleus. The perikarya of the ENK, VT, and IT neurons do not show distinct morphological differences at the level of light microscopy and are not located preferentially within the nucleus. Separate ENK, VT and IT fibers course laterally and ventrally through the hypothalamus as they descend toward the pituitary gland. All three fiber types form terminals around blood vessels in the neurohypophysis.

Histological observation of some of the endocrine glands in the sterile carp-funa hybrid (F1), with special reference to the hypophysis.

Some endocrine glands of the carp-funa hybrids were studied with a light microscope to elucidate their detailed structure and the possible causal factor of sterility in the males. Adult specimens of carp (Cyprinus carpio), gengoroh-buna (Carassius auratus cuvieri), and their hybrid (F1) were examined. The hybrid males are sterile as manifested by the failure of meiosis and seminomatous neoplasm in their testes. The hybrid females revealed well-developed ovaries, but their fertility was not tested. The hybrid hypophysis shows an intermediate condition between the parent species in the grade of ramification of the pars nervosa into the pars intermedia. Among seven types of granular cells demonstrated in the adenohypophysis, certain degenerative and anomalous changes are recognized only in the gonadotrophs of the hybrid hypophysis, especially in the female. These changes are discussed as a possible cause of sterility. A considerable amount of aldehyde fuchsin stainable neurosecretory material occurs in the cells of the nucleus preopticus and in the pars nervosa. The nucleus lateralis tuberis exhibits a histologically healthy condition.

Topography of estrogen target cells in the forebrain of goldfish, Carassius auratus.

The topographical distribution of estradiol-concentrating cells in the brain of male and female teleost, Carassius auratus, was determined by autoradiography, after injection of 3H estradiol-17beta. Radioactively labeled neurons are found in specific regions of the forebrain, with a similar topographical distribution both in males and females. Regions of accumulation of estrogen target cells include the supracommissural area of the telencephalon, the preoptic area, the central hypothalamic area and the thalamic area. Unlabeled estradiol injected prior to the administration of 3H estradiol reduces or eliminates nuclear uptake of radioactivity. The autoradiographic results demonstrate the existence of estrogen target neurons in the teleost forebrain at sites similar to those described in mammalian brains. However, under the conditions of the experiment, there exists no pallial representation of estrogen target cells.

[Interrelationships of the neurosecretory cells and capillaries in the preoptic nucleus of the carp]. / O vzaimootnosheniiakh neirosekretornykh kletok s kapilliarami v preopticheskom iadre karpa.

Three layers are tentatively recognized in the wall of a blood capillary of nucleus praeopticus (NP) of the carp, Cyprinus carpio L: the inner layer--endothelium, the middle layer--basement membrane with pericytes, and the outer broken layer containing mostly leucocytes and fibroblast-like cells. 1). The plasmalemma of the neurosecretory cells (NSC) makes a direct contact with a capillary: (a) with the plasmalemma of a sole-like process of the endothelial cell (0.1%), (b) with the basement membrane of the capillary (4.9%), (c) with the plasmalemma of the outer layer cells (41.3%). 2) The plasmalemma of the NSC is separated from the capillary by a glial process (53.7%). The first kind of the contact is frequent in the ventral part (3.7%), and the second one--in the dorsal part (29.4%) of NP. The different relations of the NSC--the capillary of the ventral and dorsal parts are due to heterochronism of their development in ontogenesis.

The connections of the trigeminal and facial motor nuclei in the brain of the carp (Cyprinus carpio L.) as revealed by anterograde and retrograde transport of horseradish peroxidase.

The connections of the rostral and caudal parts of the trigeminal and facial motor nuclei in the carp were studied with the horseradish peroxidase technique. Following ionophoretic peroxidase injections in these motor nuclei, retrogradely labeled cells were observed together with anterogradely labeled motor cell processes. Several cellular areas in thalamus, cerebellum and medulla oblongata were shown to project to the V and VII motor nuclei. Labeled cells were found in the inferior lobe and the glomerular complex of the thalamus. In the medulla oblongata, cells in the descending trigeminal nucleus, reticular nuclei and motor nuclei other than those injected were labeled. Besides these conspicuous projections several smaller connections were also found. These findings are discussed on their significance to respiratory function. Anterogradely labeled cellular processes constitute a relatively simple network of fiber connections between the various motor nuclei and the reticular nuclei of the brainstem. This apparently dendritic system of the bulbar motor complex shows a certain degree of similarity to the structure of the motor system in the spinal cord, and might play a role in the coordinated control of the muscular system.

[Electron microscopic study of the peptidergic neurosecretory cells of the preoptic nucleus of carp]. / Elektronnomikroskopicheskoe issledovanie peptidergicheskikh neirosekretornykh kletok preopticheskogo iadra karpa.

The preoptic neurosecretory cells (NSC) in sexually mature female Cyprinus carpio L. have been studied with electron microscope in winter. Separate weakly differentiated NSC and differentiated NSC of small and mean size are localized in the ventral part of Nucleus preopticus. The big highly differentiated NSC are mostly localized in the dorsal part of Nucleus preopticus. Among these types of the cells are established the light and dark forms. The "picnomorphic" cells are found only in the dorsal part of Nucleus preopticus. The light NSC are subdivided into two following kinds of cells: 1. NSC with a great amount of moderately dilated channels of the rough endoplasmic reticulum and scarce neurosecretory granules; 2. NSC with a small amount of narrow channels of the rough endoplasmic reticulum and many neurosecretory granules. The various types of differentiated NSC reflect the asynchronous formation of neurosecretory granules. The Nucleus preopticus of C. carpio has mostly the light highly differentiated NSC. It is supposed that most of the NSC in Nucleus preopticus produce and storage neurosecretory granules.