Kisspeptin (product of the kiss1 gene) is the most potent known activator of the hypothalamo-pituitary-gonadal axis. Both kiss1 and the kisspeptin receptor are highly expressed in the hypothalamus of vertebrates, and low doses of kisspeptin have a robust and long-lasting stimulatory effect on the rate of action potential firing of hypophysiotropic gonadotropin releasing hormone-1 (GnRH1) neurons in mice. Fish have multiple populations of GnRH neurons distinguished by their location in the brain and the GnRH gene that they express. GnRH3 neurons located in the terminal nerve (TN) associated with the olfactory bulb are neuromodulatory and do not play a direct role in regulating pituitary-gonadal function. In medaka fish, the electrical activity of TN-GnRH3 neurons is modulated by visual cues from conspecifics, and is thought to act as a transmitter of information from the external environment to the central nervous system. TN-GnRH3 neurons also play a role in sexual motivation and arousal states, making them an important population of neurons to study for understanding coordination of complex behaviors. We investigated the role of kisspeptin in regulating electrical activity of TN-GnRH3 neurons in adult medaka. Using electrophysiology in an intact brain preparation, we show that a relatively brief treatment with 100 nM of kisspeptin had a long-lasting stimulatory effect on the electrical activity of an extrahypothalamic population of GnRH neurons. Dose-response analysis suggests a relatively narrow activational range of this neuropeptide. Further, blocking action potential firing with tetrodotoxin and blocking synaptic transmission with a low Ca(2+)/high Mg(2+) solution inhibited the stimulatory action of kisspeptin on electrical activity, indicating that kisspeptin is acting indirectly through synaptic regulation to excite TN-GnRH3 neurons. Our findings provide a new perspective on kisspeptin's broader functions within the central nervous system, through its regulation of an extrahypothalamic population of GnRH neurons involved in multiple neuromodulatory functions.

Given that novice drivers tend to be young, and teenagers and young adult drivers are involved in the greatest number of accidents, it is important that we understand what factors impact the driving skills of this population of drivers. The primary aim of the present study was to understand the impact of gender, organized athletics, and video gaming on driving skills of novice drivers under real-world driving conditions. Novice driving students having less than five hours driving experience previous to a normal driving lesson were evaluated on their self-confidence (self-reported) prior to the lesson and driving skill evaluated by their instructor during the course of the lesson. Information was collected about gender, age, whether or not the students were involved in organized athletics, and the extent of their video game playing. There was no impact of gender or extent of video game playing on driving skills. Females were significantly less self-confident with driving than males, but this did not translate to gender differences in driving skills. Being involved in organized athletics-either currently or in the past-significantly enhanced driving skills in both females and males. Finally, novice drivers' age was negatively correlated with driving skills. That is, younger novice drivers (especially males) had better driving skills than older novice drivers. This is counter to popular belief that young drivers lack technical driving skills because they have less experience behind the wheel. Based on the results of the current study, we hypothesize that the relatively high accident rate of younger drivers (especially male drivers) is most likely due to inattention to safety considerations rather than lack of technical driving ability.

Understanding development of gonadotropin-releasing hormone (GnRH) neuronal circuits is fundamental to our understanding of reproduction, but not yet well understood. Most studies have been focused on GnRH neurons located in the hypothalamus and preoptic area (POA), which directly regulate the pituitary-gonadal axis. In zebrafish (Danio rerio), two forms of GnRH have been identified: GnRH2 and GnRH3. GnRH3 neurons in this species plays two roles: hypophysiotropic and neuromodulatory, depending on their location. GnRH3 neurons in the ventral telencephalon, POA, and hypothalamus control pituitary-gonadal function; in other areas (e.g., terminal nerve), they are neuromodulatory and without direct action on reproduction. To investigate the biology of GnRH neurons, a stable line of transgenic zebrafish was generated in which the GnRH3 promoter drives expression of a bright variant of green fluorescent protein (Emerald GFP, or EMD). This provides unprecedented sensitivity in detecting and imaging GnRH3 neurons during early embryogenesis in the transparent embryo. Using timelapse confocal imaging to monitor the time course of GnRH3:EMD expression in the live embryo, we describe the emergence and development of GnRH3 neurons in the olfactory region, hypothalamus, POA, and trigeminal ganglion. By 50 h post fertilization, these diverse groups of GnRH3 neurons project broadly in the central and peripheral nervous systems and make anatomical connections with each other. Immunohistochemistry of synaptic vesicle protein 2 (a marker of synaptic transmission) in this transgenic model suggests synaptic formation is occurring during early development of the GnRH3 neural network. Electrophysiology reveals early emergence of responsiveness to the stimulatory effects of kisspeptin in terminal nerve GnRH3 neurons. Overall, our findings reveal that the GnRH3 neuronal system is comprised of multiple populations of neurons as a complicated network.

The University of California at Los Angeles (UCLA) quickly and comprehensively transformed its laboratory safety program following a chemistry accident in December 2008 that caused the death of a researcher. UCLA's Chancellor immediately announced sweeping changes in the laboratory safety program and challenged UCLA to become "Best in Class" in academic laboratory safety. Given the size of UCLA's lab researcher population and the decentralized nature of the campus, it became a daunting task to reach out and improve compliance with newly implemented safety policies. The Office of Environment, Health and Safety (EH&S) improved their operations, enhanced overall inspection procedures, and instituted mandatory laboratory safety training of PIs and researchers. A Laboratory Hazard Assessment Tool (LHAT) was implemented to aid in the identification of hazards, track laboratory space and personnel, and guide lab groups in compliance with personal protective equipment (PPE) policies. Laboratory inspections were increased by more than 4-fold in 2012 as compared to 2007 and now followed a comprehensive checklist to improve reproducibility and thoroughness of the inspection process. To provide better customer service to the research community and expedite corrective actions, all laboratory inspection reports are required to be issued the next business day. Unannounced PPE inspections began in 2010, and inspection findings showed a robust decrease in non-compliance with the PPE policy within one year of its implementation. As of the first half of 2013, all PIs of active laboratory research programs had completed the initial, in-class PI-specific laboratory safety training. In 2012, more than 20,000 EH&S safety classes were completed either in-class or online by PIs, lab supervisors, research staff, students, and visitors to UCLA's research laboratories. Overall, analysis of outcomes from changes in UCLA's lab safety program indicates rapid improvements in compliance with lab safety regulations such as increased PPE use. Lab safety changes required commitment and cooperation at all levels - from executive leadership, to health and safety experts, to deans and department chairs, and to PIs and researchers. © 2014 The Authors.

GnRH neurons integrate internal and external cues to control sexual maturation and fertility. Homeostasis of energy balance and food intake correlates strongly with the status of reproduction. Neuropeptides secreted by the hypothalamus involved in modulating energy balance and feeding may play additional roles in the regulation of reproduction. Hypocretin (Hcrt) (also known as orexin) is one such peptide, primarily controlling sleep/wakefulness, food intake, and reward processing. There is a growing body of evidence indicating that Hcrt/orexin (Hcrt) modulates reproduction through interacting with the hypothalamo-pituitary-gonadal axis in mammals. To explore potential morphological and functional interactions between the GnRH and Hcrt neuronal systems, we employed a variety of experimental approaches including confocal imaging, immunohistochemistry, and electrophysiology in transgenic zebrafish, in which fluorescent proteins are genetically expressed in GnRH3 and Hcrt neurons. Our imaging data revealed close apposition and direct connection between GnRH3 and Hcrt neuronal systems in the hypothalamus during larval development through adulthood. Furthermore, the Hcrt receptor (HcrtR) is expressed in GnRH3 neurons. Electrophysiological data revealed a reversible inhibitory effect of Hcrt on GnRH3 neuron electrical activity, which was blocked by the HcrtR antagonist almorexant. In addition, Hcrt had no effect on the electrical activity of GnRH3 neurons in the HcrtR null mutant zebrafish (HcrtR^-/-). Our findings demonstrate a close anatomical and functional relationship between Hcrt and GnRH neuronal systems in zebrafish. It is the first demonstration of a link between neuronal circuits controlling sleeping/arousal/feeding and reproduction in zebrafish, an important animal model for investigating the molecular genetics of development.

Bisphenol A (BPA) is a well-known environmental, endocrine-disrupting chemical, and bisphenol S (BPS) has been considered a safer alternative for BPA-free products. The present study aims to evaluate the impact of BPA and BPS on the reproductive neuroendocrine system during zebrafish embryonic and larval development and to explore potential mechanisms of action associated with estrogen receptor (ER), thyroid hormone receptor (THR), and enzyme aromatase (AROM) pathways. Environmentally relevant, low levels of BPA exposure during development led to advanced hatching time, increased numbers of GnRH3 neurons in both terminal nerve and hypothalamus, increased expression of reproduction-related genes (kiss1, kiss1r, gnrh3, lhβ, fshβ, and erα), and a marker for synaptic transmission (sv2). Low levels of BPS exposure led to similar effects: increased numbers of hypothalamic GnRH3 neurons and increased expression of kiss1, gnrh3, and erα. Antagonists of ER, THRs, and AROM blocked many of the effects of BPA and BPS on reproduction-related gene expression, providing evidence that those three pathways mediate the actions of BPA and BPS on the reproductive neuroendocrine system. This study demonstrates that alternatives to BPA used in the manufacture of BPA-free products are not necessarily safer. Furthermore, this is the first study to describe the impact of low-level BPA and BPS exposure on the Kiss/Kiss receptor system during development. It is also the first report of multiple cellular pathways (ERα, THRs, and AROM) mediating the effects of BPA and BPS during embryonic development in any species.

Kisspeptin1 (product of the Kiss1 gene) is the key neuropeptide that gates puberty and maintains fertility by regulating the gonadotropin-releasing hormone (GnRH) neuronal system in mammals. Inactivating mutations in Kiss1 and the kisspeptin receptor (GPR54/Kiss1r) are associated with pubertal failure and infertility. Kiss2, a paralogous gene for kiss1, has been recently identified in several vertebrates including zebrafish. Using our transgenic zebrafish model system in which the GnRH3 promoter drives expression of emerald green fluorescent protein, we investigated the effects of kisspeptins on development of the GnRH neuronal system during embryogenesis and on electrical activity during adulthood. Quantitative PCR showed detectable levels of kiss1 and kiss2 mRNA by 1 day post fertilization, increasing throughout embryonic and larval development. Early treatment with Kiss1 or Kiss2 showed that both kisspeptins stimulated proliferation of trigeminal GnRH3 neurons located in the peripheral nervous system. However, only Kiss1, but not Kiss2, stimulated proliferation of terminal nerve and hypothalamic populations of GnRH3 neurons in the central nervous system. Immunohistochemical analysis of synaptic vesicle protein 2 suggested that Kiss1, but not Kiss2, increased synaptic contacts on the cell body and along the terminal nerve-GnRH3 neuronal processes during embryogenesis. In intact brain of adult zebrafish, whole-cell patch clamp recordings of GnRH3 neurons from the preoptic area and hypothalamus revealed opposite effects of Kiss1 and Kiss2 on spontaneous action potential firing frequency and membrane potential. Kiss1 increased spike frequency and depolarized membrane potential, whereas Kiss2 suppressed spike frequency and hyperpolarized membrane potential. We conclude that in zebrafish, Kiss1 is the primary stimulator of GnRH3 neuronal development in the embryo and an activator of stimulating hypophysiotropic neuron activities in the adult, while Kiss2 plays an additional role in stimulating embryonic development of the trigeminal neuronal population, but is an RFamide that inhibits electrical activity of hypophysiotropic GnRH3 neurons in the adult.