Epicortical slow potential shifts and sensory-evoked potentials are related to seizure propensity in gerbils

Gerbils were assessed for behavioural tendency by scoring seizure severity and the amount of ambulatory and rearing activities in a novel `open-field' arena. Seizure-prone animals exhibited seizures on early open-field trials (1–2) and later performed more ambulatory activity than non-seizure-prone animals. Two weeks later, two groups of both seizure prone and non-seizure prone animals were chronically implanted with six silver/silver chloride ball electrodes for recordings during behaviour. Electrodes were on the surfaces of the frontal, parietal and occipital cortices bilaterally. In one group these were used to record slow potential shifts; in the other, visual- and acoustic-evoked responses. Larger negative and positive slow shifts occurred in seizure- prone animals. Most evident were the larger positive right frontal shifts and negative left occipital shifts. Seizure tendency was related to the amplitude of these waveforms. Visual-evoked potential amplitudes were generally larger and latencies shorter in seizure-prone animals, especially in the right occipital and left parietal cortices. Seizure susceptibility was associated with increased visual-evoked potential amplitude in the right frontal and left occipital cortices, and with reduced latency of both auditory-and visual-evoked responses in the left occipital cortex. The discussion highlights a role for glia in slow shift generation and the association of large shifts with enhanced sensory-evoked responses, especially in seizure-prone animals. Accepted: 21 January 1998     

Swimming response of goldfish,Carassius auratus,and the tetra,Hemigrammus caudovittatus,larvae to individual food items and patches

Synopsis Swimming speed and swimming path of goldfish and tetra larvae were studied in aquaria containing food patches composed of decapsulated cysts and immobilized nauplii of Artemia salina or sparsely distributed prey. The mean swimming speed of starved larvae in the medium without food was about four times higher than the speed of larvae feeding in a patch. Satiated larvae swam about 1.5 times slower than hungry fish. Consumption of single prey items by starved larvae caused the following sequence of swimming responses: handling pause (cessation of swimming), slow swimming in a restricted area, and fast swimming (approximately twice as fast as hungry larvae before encountering food) accompanied by a widening of the area searched (area increased searching). Mean swimming speed was constant over a broad range (10¹–10³ ind·1^–1 of food density, although at extreme (high or low) values of food density it depended on swimming responses of the predator. Frequency of visits to the different parts of the aquarium strongly depended on encounters of hungry fish with food particles or patches.     

Sustained potential shifts and changes in acoustic evoked potentials after presentation of a non-acoustic priming stimulus to carp (Cyprinus carpio).

1. Recordings were made from the region of the midbrain tectum and torus semicircularis of sustained potential shifts (SPS) to a non-acoustic priming stimulus and the change in subsequent acoustic evoked potentials (AEPs) to a train of six clicks after a long rest. 2. In the absence of priming stimuli (a jet of saline or water to the flank) the AEP to the first click in a train had the highest amplitude; with these stimuli it became the most attenuated. 3. The SPS to both non-acoustic stimuli was initially (ca 4 sec) negative, then became positive for a similar time period. 4. After saline jet the tectal and the torus AEP amplitude was significantly correlated with the torus SPS; after water jet, the tectal and the torus AEP durations were correlated with the SPS. 5. Application of alumina gel to the posterior telencephalic border caused elevation of the torus AEP amplitude after some 5 hr.     

New observations on the fine structure of the liver in goldfish (Carassius auratus)

Summary A re-examination of goldfish liver was made through the use of SEM of fractured samples and TEM of ultrathin-sections and freeze-etch replicas. Several new hepatic fine structures described in the present study are morphologically similar to those reported previously in many higher vertebrates including mammals. Hepatic sinusoids of goldfish contain fenestrations which are arranged into sieve plates. Although the hepatic plates are made up of two layers of hepatocytes, the parenchymal cells of goldfish liver are morphologically similar to mammalian hepatocytes, particularly with respect to the sinusoidal surfaces which are studded with numerous microvilli. The intercellular surfaces of hepatocytes have both nexus and desmosomal junctions, similar to those found in various epithelial cells of higher vertebrates, as cell attachments and communication foci. Tight junctions are found mainly between the openings of the intracellular bile canaliculi and the intralobular bile ductules which are situated in the center of the bicellular hepatic plate.Supported in part by Grants # GM92 and ES07017     

Effects of exposure to sublethal levels of cadmium upon water-electrolyte status in the goldfish (Carassius auratus)

Goldfish were exposed to sublethal levels of cadmium (means of 44.5 and 380 μg Cd⁺⁺/l) for periods of 25 and 50 days, and their water-electrolyte status evaluated by reference to plasma and muscle levels of sodium, potassium and chloride and muscle water content. Significant changes in plasma chloride, tissue potassium and tissue water content were observed after 25 days in both test solutions. Specimens held at the more dilute cadmium concentration were apparently able to compensate for most of the initial cadmium effect and, after 50 days exposure, were characterized only by a continuing depression in plasma sodium level. This suggests that the cadmium MATC value for this species under the conditions employed is probably less than 45 μg Cd⁺⁺/l. Goldfish exposed to 380 μg Cd⁺⁺/l for 50 days exhibited significant deviations in plasma sodium and chloride levels as well as in tissue sodium and water content, and these parameters may provide useful indices of cadmium effects at sublethal concentrations.     

X-ray microanalysis of myotomal muscle in the sunbleak, Leucaspius delineatus (Heckel) and goldfish, Carassius auratus gibelio

The functioning of a group of muscle fibres as a tissue that performs a well characterized type of contraction (slow or fast) depends on their biochemical and structural organization that is already well established. The biochemical and structural diversities between three types of fish muscle fibres found also a reflection in the content of light elements. The present work demonstrates significant differences in the content of diffusible elements (Cl, K, Na, and Mg) and bound elements (P and S) between the muscle fibres types. In general all muscle fibre types of goldfish (Carassius auratus gibelio) that belongs to stationary slow-swimming fish has lower K/Na ratios than those in all three fibre types of fast swimming sunbleak (Leucaspius delineatus).     

Morphological identification of live gonadotropin,growth-hormone,and prolactin cells in goldfish (Carassius auratus) pituitary-cell cultures

To better understand neuroendocrine regulation and the intracellular mechanisms mediating pituitary-hormone release, it is necessary to study the physiology of identified single cells. We have developed a system to identify gonadotropin, growth-hormone, and prolactin cells in primary cultures of goldfish pituitary cells. Using Nomarski differential interference-contrast microscopy, the unique morphologies of discrete subpopulations of cells were characterized. To aid in the initial characterization of different pituitary-cell types, a discontinuous Percoll density-gradient cell-separation technique was developed. This method provided fractions enriched with functional gonadotropin, growth-hormone, and prolactin cells. The morphology of each cell type was initially characterized in enriched fractions of immunofluorescently labelled cells using differential interference-contrast microscopy. The cell type-specific morphologies were then confirmed in live pituitary-cell cultures. Gonadotropin, growth-hormone, and prolactin cells were correctly identified in live pituitary-cell cultures. Gonadotropin, growth-hormone, and prolactin cells were correctly identified in live mixed cultures in 92, 94, and 100% of the trials, respectively. The ability to directly identify cells in primary cultures allows the physiological study of identified single cells with minimal pretreatment.     

Pheromone discrimination ability of olfactory bulb mitral and ruffed cells in the goldfish (Carassius auratus)

Significant anatomical differences characterizing mitral cells and ruffed cells were published by Kosaka and Hama in three teleost species. Physiological responses from both types of relay neurons have now been recorded extracellularly and simultaneously in the plexiform layer using a single tungsten microelectrode. During interstimulus intervals mitral cells responded with higher, frequently burst-like impulse rates triggered by the activity of epithelial receptor neurons. Ruffed cell impulse rates were low, and each action potential triggered a long-lasting, continuously variable, integrated granule cell potential. During olfactory stimulation with important biological stimuli such as preovulatory and ovulatory pheromones, a probable alarm pheromone and amino acids contrasting interactions between mitral cells and ruffed cells resulting in a drastic intensification of centrally transmitted information were frequently recorded. Individual neurons excellently discriminated stimuli. Irrespective of the physiological relevance of stimuli, however, similarities were recorded in the distribution of excitatory, inhibitory and indifferent responses.     

The interactive effects of hypoxemia, hyperoxia, and temperature on the gill morphology of goldfish (Carassius auratus)

Acclimation of crucian carp and goldfish to temperatures below 15°C causes covering of the gill lamellae by a mass of cells termed the interlamellar cell mass (ILCM). Here we explore the cues underlying gill remodeling (removal or growth of an ILCM) and specifically test the hypotheses that 1) depletion of internal O(2) stores in the absence of any change in external O(2) status can trigger the removal of the ILCM in goldfish acclimated to 7°C, 2) exposing fish acclimated to 25°C to an abundance of O(2) (hyperoxia) can reverse the gill remodeling (i.e., cause the covering of lamellae by an expansion of the ILCM), and 3) neuroepithelial cells (NECs) are involved in signaling the shedding of the ILCM. Hypoxemia induced by phenylhydrazine (anemia) or 5% CO caused a decrease in the ILCM from 80% to 23% and 35%, respectively. Hyperoxia exposure at 25°C caused an increase to 67% of total ILCM and a smaller decrease in the size of the ILCM when fish were transferred from 7 to 25°C. Daily sodium cyanide injections were used to stimulate NECs; this treatment led to a significant decrease in the ILCM. Thus, the three major conclusions of this study are 1) that gill remodeling can occur during periods of internal hypoxemia, 2) that O(2) supply and demand may be a significant driving force shaping gill remodeling in goldfish, and 3) the NECs may play a role in triggering the shedding of the ILCM during hypoxia.     

The variability of the reaction to a visual stimulus in goldfish, Carassius auratus L. (Cyprinidae: Pisces)

Animal's responses to repeated identical stimulation are notoriously variable. The variability is not always resulted from differences between individuals, stimulus action, or habituation. Goldfish's responses to a visual stimulus were studied in order to reveal the nature of variability. The experimental device consisted of a white circular corridor, and the black band stimulus drown across the corridor. Fish responded to the stimulation by an increase in frequency of turns in the vicinity of the band. The response, however, was not always positive: sometimes fish passed by the band with no turns (negative response). The significant tendency to repeat positive and negative responses by series was observed. Intensity of the response differed between individuals, and some individuals never responded at all. The simulating dynamic model of goldfish behaviour was studied both qualitatively and quantitatively. The model assumes that the control mechanism of the behaviour is influenced by the noise (fluctuations of nervous activity) which causes phase transitions in the mechanism. The similarity of the model's behaviour to experimental data suggests that the response variability observed in goldfish resulted from the self-organising process. Therefore, the variability should persist even in the absence of any variations in an external stimulation, as well as in the absence of individual differences.     

Influence of forskolin and carbachol on intestinal absorption of horseradish peroxidase in the goldfish (Carassius auratus)

The transepithelial route for mucosa-to-serosa transport of the tracer macromolecule horseradish peroxidase (HRP; MW 40 kDa) and modulation of this transport by forskolin and carbachol have been studied in vi-tro in stripped goldfish intestinal epithelium mounted in Ussing-type chambers. Uptake and transport have been investigated by measuring the HRP flux from the muco-sal to serosal sides by an enzymatic method and by visualising HRP reaction products in the mucosa with electron-microscopical techniques. Both the cholinergic agonist carbachol (which is thought to increase intracellular Ca²⁺ and activate protein kinase C activity) and forskolin (a direct activator of adenylylcyclase) affect the amount of enzymatically active HRP in the tissue. In control tissue, HRP product is found only within the epithelial cells, the transepithelial flux reaching a constant value of about 1.5 pmoles/cm² per h. Carbachol increases the amount of HRP product in the cells, but has no significant effect on the HRP flux compared with control values. Forskolin decreases the amount of HRP product in the cells; however, in the presence of forskolin, the lateral intercellular spaces become filled with HRP product. HRP is found in the lamina propria and the transepithelial protein flux increases more than 2.5-fold. In the presence of forskolin plus carbachol, the results are no different from the control. It is concluded that carbachol increases the endocytotic uptake of HRP, whereas forskolin inhibits the uptake but increases the paracellular permeability for HRP in goldfish intestine. Received: 10 July 1995 / Accepted: 4 February 1996     

Interactions between gonadotropin-releasing hormone (GnRH) and orexin in the regulation of feeding and reproduction in goldfish (Carassius auratus)

Links between energy homeostasis and reproduction have been demonstrated in vertebrates. As a general rule, abundant food resources favor reproduction whereas low food availability induces an inhibition of reproductive processes. In both mammals and fish, gonadotropin-releasing hormone (GnRH) and orexin (OX) are hypothalamic neuropeptides that play critical roles in the regulation of sexual behavior and appetite, respectively. In order to assess possible interactions between orexin and GnRH in the control of feeding and reproduction in goldfish, we examined the effects of chicken GnRH (cGnRH-II) intracerebroventricular (ICV) injection on feeding behavior and OX brain mRNA expression as well as the effects of orexin ICV injections on spawning behavior and cGnRH-II brain mRNA expression. Treatment with cGnRH-II at doses that stimulate spawning (0.5 ng/g or 1 ng/g) resulted in a decrease in both food intake and hypothalamic orexin mRNA expression. Treatment with orexin A at doses that stimulate feeding (10 ng/g) induced an inhibition of spawning behavior and a decrease in cGnRH-II expression in the hypothalamus and optic tectum-thalamus. Our results suggest that the anorexigenic actions of cGnRH-II in goldfish might be in part mediated by OX and that orexin inhibits reproductive behavior in part via the inhibition of the GnRH system. Our data suggest the existence of a coordinated control of feeding and reproduction by the orexin and GnRH systems in goldfish.     

Investigation of de novo cholesterol synthetic capacity in the gonads of goldfish (Carassius auratus) exposed to the phytosterol beta-sitosterol

Total and intra-mitochondrial gonadal cholesterol concentrations are decreased in fish exposed to the phytoestrogen beta-sitosterol (beta-sit). The present study examined the potential for beta-sit to disrupt de novo cholesterol synthesis in the gonads of goldfish exposed to 200 microgram/g beta-sit and 10 microgram/g 17beta-estradiol (E2; estrogenic control) by intra-peritoneal Silastic^® implants for 21 days. The de novo cholesterol synthetic capacity was estimated by incubating gonadal tissue with 14C-acetate for a period of 18 hours, followed by chloroform/methanol lipid extraction and thin layer chromatography (TLC) lipid separation. Lipid classes were confirmed using infrared spectroscopy. Plasma testosterone (T) and total cholesterol concentration were measured and gonadosomatic index (GSI) was calculated. Plasma T was significantly reduced in male beta-sit-treated fish compared to control and E2-treated fish (p < 0.001). 14C-Acetate incorporation into cholesterol and cholesterol esters was not significantly different among treatment groups for male and female fish, however, 14C-enrichment was higher than expected in both triglycerides (TG) and free fatty acids (FFA). FFA incorporation was significantly higher in male control fish than either beta-sit or E2 treatments (p = 0.005). Plasma cholesterol concentration was significantly increased in the male beta-sit treatment group compared to controls (p = 0.027). These results indicate gonadal de novo cholesterol biosynthetic capacity is not disrupted by beta-sit or E2 treatment in early recrudescing male or female goldfish, while plasma cholesterol and steroid concentrations are sensitive to beta-sit exposure.     

Formation of viable but non-culturable state (VBNC) of Aeromonas hydrophila and its virulence in goldfish, Carassius auratus

In this study we investigated the viable but non-culturable (VBNC) state of Aeromonas hydrophila and its virulence in goldfish. Aeromonas hydrophila cultured in a 0.35% NaCl solution at pH 7.5 and at 25 degrees C for 50 days showed the VBNC state. In the VBNC state we were unable to detect viable bacteria by the plate count method but we did find 10(4) cells/ml by the direct viable count microscopical method after staining with fluorescein diacetate and ethidium bromide. The virulence comparison in goldfish showed that bacteria cultured at 25 degrees C for 1 day in a 0.35% NaCl solution were more virulent than bacteria cultured for 28 days. VBNC bacteria showed lower virulence in goldfish compared to 28-day-cultured bacteria by intraperitoneal injection. The results from the study suggest that A. hydrophila can remain in the aquatic environment for prolonged periods in the VBNC state but those cells are not pathogenic to goldfish.