Embryonic exposure to ethanol is known to affect neurochemical systems in rodents and increase alcohol drinking and related behaviors in humans and rodents. With zebrafish emerging as a powerful tool for uncovering neural mechanisms of numerous diseases and exhibiting similarities to rodents, the present report building on our rat studies examined in zebrafish the effects of embryonic ethanol exposure on hypothalamic neurogenesis, expression of orexigenic neuropeptides, and voluntary ethanol consumption and locomotor behaviors in larval and adult zebrafish, and also effects of central neuropeptide injections on these behaviors affected by ethanol. At 24 h post-fertilization, zebrafish embryos were exposed for 2 h to ethanol, at low concentrations of 0.25% and 0.5%, in the tank water. Embryonic ethanol compared to control dose-dependently increased hypothalamic neurogenesis and the proliferation and expression of the orexigenic peptides, galanin (GAL) and orexin (OX), in the anterior hypothalamus. These changes in hypothalamic peptide neurons were accompanied by an increase in voluntary consumption of 10% ethanol-gelatin and in novelty-induced locomotor and exploratory behavior in adult zebrafish and locomotor activity in larvae. After intracerebroventricular injection, these peptides compared to vehicle had specific effects on these behaviors altered by ethanol, with GAL stimulating consumption of 10% ethanol-gelatin more than plain gelatin food and OX stimulating novelty-induced locomotor behavior while increasing intake of food and ethanol equally. These results, similar to those obtained in rats, suggest that the ethanol-induced increase in genesis and expression of these hypothalamic peptide neurons contribute to the behavioral changes induced by embryonic exposure to ethanol.

Recent studies in zebrafish have shown that exposure to ethanol in tank water affects various behaviors, including locomotion, anxiety and aggression, and produces changes in brain neurotransmitters, such as serotonin and dopamine. Building on these investigations, the present study had two goals: first, to develop a method for inducing voluntary ethanol intake in individual zebrafish, which can be used as a model in future studies to examine how this behavior is affected by various manipulations, and second, to characterize the effects of this ethanol intake on different behaviors and the expression of hypothalamic orexigenic peptides, galanin (GAL) and orexin (OX), which are known in rodents to stimulate consumption of ethanol and alter behaviors associated with alcohol abuse. Thus, we first developed a new model of voluntary intake of ethanol in fish by presenting this ethanol mixed with gelatin, which they readily consume. Using this model, we found that individual zebrafish can be trained in a short period of time to consume stable levels of 10% or 20% ethanol (v/v) mixed with gelatin and that their intake of this ethanol-gelatin mixture leads to pharmacologically-relevant blood ethanol concentrations which are strongly, positively correlated with the amount ingested. Intake of this ethanol-gelatin mixture increased locomotion, reduced anxiety, and stimulated aggressive behavior, while increasing expression of GAL and OX in specific hypothalamic areas. These findings, confirming results in rats, provide a method in zebrafish for investigating with forward genetics and pharmacological techniques the role of different brain mechanisms in controlling ethanol intake.

Exercise-induced vascular endothelial adaptations in the kidney are not well understood. Therefore, we investigated the impact of voluntary wheel running (VWR) on the abundance of endothelial nitric oxide synthase (eNOS) and extracellular superoxide dismutase (EC SOD), in kidney and lung, and other SOD isoforms and total antioxidant capacity (TAC), in kidney. We also determined whether VWR influences susceptibility to acute kidney injury (AKI). Male Sprague-Dawley and Fisher 344 rats, VWR or sedentary for 12 weeks, were subjected to AKI (uninephrectomy (UNX) and 35 min of left kidney ischaemia-24 h reperfusion, IR). We measured glomerular filtration rate (GFR) and renal plasma flow (RPF), and analysed renal structural injury. Running was comparable between strains and VWR reduced body weight. In Sprague-Dawley rats, VWR reduced eNOS and EC SOD, but increased Mn SOD in kidney. Similar changes were seen after 6 weeks of VWR in Sprague-Dawley rats. In Fisher 344 rats, VWR increased eNOS, all SOD isoforms and TAC in kidney. Both strains increased eNOS and EC SOD in lung with VWR. Compared to UNX alone, UNX-IR injury markedly reduced renal function for both strains; however, in the Sprague-Dawley rats, VWR exacerbated falls in GFR and RPF due to UNX-IR, whereas in the Fisher 344 rats, GFR was unaffected by VWR. Some indices of renal structural injury due to UNX-IR tended to be worse in SD vs. F344. Our study demonstrates that genetic background influences the effect of exercise on kidney eNOS and EC SOD, which in turn influence the susceptibility to AKI.

One mechanism by which the vasculature benefits from exercise (EX) is by increases in blood flow (BF). This causes increases in shear stress and induction of the endothelial nitric oxide synthase (eNOS) and antioxidant, extracellular superoxide dismutase (EC SOD). The kidney, however, undergoes reduced BF during EX and therefore may be at risk for falls in NO and antioxidant bioavailability. Genetic differences may also dictate this response. Therefore, we investigated kidney cortex (KC) responses to EX in the Sprague Dawley (SD) and Fisher 344 (F344) rat. Male SD and F344 rats (10–12 wks old) voluntarily wheel ran for 12 wks (24 hr access) or remained sedentary (SED). Running activity was similar between strains. Nitrate/nitrite (NOx) levels and eNOS and EC SOD abundance in the KC of F344 rats increased with EX (Table; *p≤0.05 vs. SED). In contrast, SD rats decreased their renal eNOS and EC SOD with no change in KC NOx. The oxidative stress marker p22phox also increased in the KC of EX F344 but decreased in the EX SD rat. Another oxidative stress marker, H2O2, measured in KC, increased only in EX F344 rats. These findings indicate that a strain difference exists in the response to 12 wks voluntary EX between the F344 and SD rat. F344 rats increased their KC eNOS and EC SOD and this was interestingly also associated with increased oxidative stress. Conversely, SD rats had the opposite response to EX in their KC: reduced eNOS and EC SOD and either decreased p22phox or no change in H2O2. Mechanisms responsible for these marked differences in renal responses to EX remain to be determined. eNOS NOx EC SOD p22phox H2O2 n Arbitrary Units nmol/mg Arbitrary Units Arbitrary Units nmol/mg F344 SED 12 6±1.2 8±1.7 68±23 5±0.5 0.2±0.04 EX 12 18±3.2* 15±6.3* 122±27* 8±0.6* 0.6±0.14* SD SED 6 64±2.6 9±0.4 13±1.0 9±0.4 0.4±0.07 EX 6 56±2.2* 10±0.4 8±0.7* 7±0.6* 0.5±0.07