In the early twentieth century, Karl von Frisch performed seminal work on the organization of social behavior of honey bees. Much of his work involved training individual foragers to distant artificial feeders. Similar training methods have been used in research laboratories for the better part of a century, and these methods lend themselves well to advanced undergraduate biology classes in animal behavior. In recent years, students have used these methods in group projects to study color preference and time-memory. In this Technical Paper, we describe the basic steps of training honey bees to a distant feeder. We also provide alternative methods for answering specific types of questions that students in animal behavior classes might wish to address.

<p dir="ltr">Stocking density, space availability, and lameness are important aspects affecting broiler chicken behavior and welfare. Stocking density refers to the weight of broiler chickens per a set area of space typically measured as kg/m2. Space availability is the amount of space per individual broiler chicken typically measured as m2/bird. Stocking density and space availability can contribute to lameness and other aspects of welfare such as footpad dermatitis, hock burn, and feather cleanliness. The behavior of broiler chickens can also be modified by stocking density, space availability, and lameness. All of these aspects are typically related to a change in activity levels which could be used as an indicator of animal welfare. To date, the majority of research that has examined the walking ability of broiler chickens has assessed how stocking density influences the development of gait problems when applied during the grower phase. However, not all broilers develop gait problems at the same point in time and it is unknown whether broilers that initially have sound gait develop gait problems at a similar rate to broilers that are initially classified as having affected gait. Further, the influence of stocking density on the progression of gait abnormalities of broilers with sound and poor gait is unknown. Finally, since space becomes more and more limited as broiler chickens increase in body weight and age, it is unknown how the provision of space during the finisher phase, when broiler chickens are gaining weight rapidly, can influence their walking ability and welfare outcomes. Stocking density is calculated based on projected final weights of broiler chickens from the time they are placed on a commercial farm, and that projected stocking density remains the same from the chick placement date. As stocking densities are increased, there is also an increase in the prevalence of lameness. Age is also known to be related to walking ability, and as broiler chickens age, there is an increase in the prevalence of lameness. While it is known that increasing stocking density and aging are both contributing factors to broiler chicken lameness, there is no previous research on if reducing stocking density at a later age can help alleviate the prevalence of lameness. To address this knowledge gap, two studies were conducted. In the first study, 784 mixed-sex Ross 708 broiler chickens in commercial barns were placed into of four treatment groups. · SOUND: Consisted of broiler chickens that were considered to have sound gait (scores of 0 and 1) and the broilers were housed at farm stocking density (6lb/ft2, 29.29 kg/m2), · AFFECTED: Consisted of broiler chickens that were considered to have affected gait (scores of 2 or higher) and were housed at farm stocking density (6lb/ft2, 29.29 kg/m2) · MIXED-F: Consisted of 50% of broiler chickens that were considered to have sound gait and 50% that were considered to have affected gait and were housed at farm stocking density (6lb/ft2, 29.29 kg/m2) · MIXED-L: Consisted of 50% of broiler chickens that were considered to have sound gait and 50% that were considered to have affected gait and were housed at half of the farm stocking density (3lb/ft2, 14.65 kg/m2) Broiler chickens were randomly selected at 33 days of age from each of four commercial barns for welfare assessments, which included gait scoring to assess walking ability, as well as the assessment of footpad dermatitis and hock burn. Broilers were then assigned to one of two gait categories based on their gait scores. Broilers were either considered to have sound gait meaning they had no or unidentifiable abnormalities, or affected gait meaning there were identifiable abnormalities. To separate treatment groups, custom-built pens (4 ft x 12 ft, 1.22 m x 3.66 m) were constructed. At 37 days of age welfare assessments were conducted again, and then the broiler chickens were placed back into the flock (Chapter 2). The behavior of the broiler chickens was recorded from the evening of day 33 to the morning of day 37 and video was analyzed using scan sampling. The proportion of broiler chickens performing target behaviors was recorded every 10 minutes in the morning (6:00 – 8:00) and evening (19:00 – 21:30). Better gait scores were observed at 37 days of age in broiler chickens in the MIXED-L group and broiler chickens in the SOUND group. The presence of hock burn was lower in broiler chickens in the SOUND group. Cleanliness scores were better for broiler chickens in the MIXED-L group and in broiler chickens in the SOUND. Stocking density impacted the proportion of broilers performing eating, drinking, sitting, and walking (P < 0.05). Walking ability impacted the proportion of broiler chickens standing, walking, and sitting (P < 0.05). To continue investigating the implementation of housing changes later in the broiler chickens’ life, a second study was conducted using 705 mixed-sex Ross 708 broiler chickens. At 7 d, broiler chickens were randomly assigned to 1 of 16 pens (46-47 birds/pen). At 28 d, half of the pens doubled in size after welfare assessments were completed (measuring 8 ft x 10 ft, 3.05 m x 2.44 m, DOUBLE), while the other half remained at the original dimensions (8 ft x 5 ft, 2.44 m x 1.5 m, SINGLE). The DOUBLE pens had an expected stocking density of 15.2 - 15.5 kg/m2 (3.11 - 3.17 lb/ft2) and an estimated space availability of 0.15 to 0.16 m2/bird while the SINGLE pens had an estimated stocking density of 30.4 - 31.1 kg/m2 (6.23-6.37 lb/ft2) and an estimated space availability of 0.07-0.08 m2/bird. Welfare assessments consisting of scoring gait, feather cleanliness and for the presence of FPD and hock burn were conducted at 22 d, 28 d, and 38 d (Chapter 4). At 38 d, broiler chickens in SINGLE pens were less likely to have a score of 0 for FPD (Wald c2 = 15.45, P < 0.0001), hock burn (Wald c2 = 7.26, P = 0.0071), and feather cleanliness (Wald c2 = 11.77, P = 0.0006) than broiler chickens in DOUBLE pens. However, broiler chickens in SINGLE pens were more likely to have a gait score of 0 compared to broiler chickens in DOUBLE pens (Wald c2 = 11.34, P = 0.0008). Broiler chicken behavior was recorded at 23-26 d (Period 1: before space increase), 28-31 d (Period 2: time of space increase), and 36-37 d (Period 3: after space increase). Behavior data were collected using focal sampling for two broiler chickens per each of the 16 pens in the morning, afternoon, and evening (Chapter 5). Broiler chickens housed in double pens had an increased frequency of leg extensions compared to broiler chickens housed in single pens (P < 0.05). Period had a significant impact on the frequency of eating, sitting, and walking and the durations of sitting, environmental pecking, standing, and walking (P < 0.05). Time of day had a significant impact on the frequency of eating, sitting, walking, preening, and leg extensions and the durations of sitting, eating, preening, and standing (P < 0.05). The interaction of age and time of day had a significant impact on the frequency of drinking and leg extensions and the durations of sitting, eating, and walking (P < 0.05). The interaction of age and treatment had a significant impact on the frequency of eating and walking and the duration of preening (P < 0.05). In conclusion, broiler chickens housed in DOUBLE pens did not exhibit a difference in behaviors compared to those in SINGLE pens, other than broilers in the DOUBLE pens performing leg extensions more often. While the first study indicated that having more space available per broiler chicken led to better walking ability, the second study showed the opposite to be true as those with more space had reduced walking ability. This indicates that changing the stocking density through manipulating space in the finisher phase may impact welfare, but further investigation is needed. Future research should first examine the effects of adding space in the finisher phase with 3 treatment groups. While the two discussed here would remain the same, the third group should start with broilers in a pen that is already the size of the DOUBLE pens and remains that way for the entire project. This will ensure that increasing space during the finisher period is beneficial rather than the additional space availability in general accounting for the differences in treatments. All treatment groups should also get fresh bedding with the pen increase to ensure the welfare measurement results are due to the changes in space availability rather than the provision of fresh litter.</p>

As zoos and aquariums have become increasingly focused on conservation education, their menageries of unique and diverse learning opportunities have been underutilized. Through a new postsecondary-level animal behavior laboratory experience at an aquarium (“ZooU”), this study demonstrates that active learning pedagogy aligned with the Next Generation Science Standards (NGSS) could facilitate expansion of education at zoos and aquariums beyond their conservation education niche. Generally, students indicated that ZooU provided new opportunities for them to explore their own interests, demonstrate their learning, and augment their previous laboratory and aquarium experiences. Following both self- and researcher assessments of the students’ work, integrated analyses revealed that students who engaged in more active learning activities at the aquarium demonstrated a greater increase in skills aligned with the NGSS. Additionally, a novel intra-individual analysis was utilized to embrace the variation between learners that typically confounds the results of education studies with repeated measures design. Common challenges for education at zoos and aquariums are discussed through the context of ZooU as a foundation for future investigations. A practical NGSS-aligned guide to field trips at zoos and aquariums—written specifically for science teachers—is also included to support broader utilization of zoos and aquariums as educational resources.

No description available.

No description available.

No description available.

No description available.

Tinnitus, ringing in the ears, is a phantom sound percept affecting roughly 10-20% of the total world population. Tinnitus severely impacts the quality of life of 10% of tinnitus sufferers, affecting their sleep, concentration, emotion, social enjoyment, and sometimes leading to depression and suicidal tendencies. In humans, most forms of tinnitus are associated with noise-exposure, leading to compensatory maladaptive plasticity of central auditory neurons. Human and animal studies suggest that tinnitus alters normal adult attentional resources. Human studies by McKenna, Hallam and Surlock 1996, suggested tinnitus-related impairment in sustained attention, vigilance, visual conceptualization and visuo-motor memory. Additionally, tinnitus may impact aspects of selective or divided attention as well as working and long-term memory. The involvement of primary auditory cortex and nicotinic signaling in selective attention, working and long-term memory has been well established. Neuronal nicotinic acetylcholine receptors (nAChRs) are present on presynaptic and postsynaptic inputs that innervate neurons across layers of primary auditory cortex (A1). Layer 5 pyramidal neurons (PNs) in the A1 are major output neurons, conveying auditory information to corticocortical and subcortical nuclei. The excitation of PNs is regulated by a complex microcircuitory of inhibitory neurons with vasointestinal peptide positive (VIP) neurons playing a key role in regulating the excitation. The focus of present studies was to: 1) Characterize tinnitus-related changes in the physiology and nAChR signaling of layer 5 PNs and VIP neurons in the A1 and 2) Determine the ability of nAChR partial/desensitizing agonists to ameliorate tinnitus pathology in subcellular studies. Wild-type, ChAT-Cre and VIP-Cre:Rosa26-loxP-stop-loxP-tdTomato (VIP-Cre:Rosa-tdTomato Long-Evans rats were used in the present study. CHAT-Cre rats allowed us to selectively express cre-inducible AAV-EF1a-DIO-hChR2(H134R)-EYFP and stimulate the cholinergic neurons of basal forebrain (BF). VIP-Cre:Rosa-tdTomato express fluorescent tdTomato protein in the VIP positive neurons allowing us to identify them under fluorescence microscopy using 550 nm wavelength. An established noise-exposure (one hour of 116 dB narrowband noise centered at 16 kHz) was used to induce behavioral tinnitus in a rat model. Approximately 50-60% noise-exposed animals (53/92) exhibited behavioral evidence of tinnitus with significant shifts in hearing threshold contiguous to the exposure frequency. Animals were classified as control, exposed tinnitus and non-tinnitus. In vitro whole-cell patch clamp recordings were performed in control and tinnitus animals. Results: Numerous tinnitus-related changes in the physiology of layer 5 PNs and VIP neurons, and changes in the activity of excitatory and inhibitory input neurons were observed. The resting membrane potential of A1 layer 5 PNs from tinnitus animals was significantly depolarized compared to PNs from unexposed controls. PNs from the A1 of animals with behavioral evidence of tinnitus showed increases in the frequency of excitatory and decreases in frequency of inhibitory spontaneous postsynaptic currents, which directly correlated with the rat’s tinnitus score. Optical stimulation of thalamocortical terminals from PNs in tinnitus animals evoked significantly larger excitatory/inward currents than in currents evoked in PNs from controls. A1 layer 5 PNs showed tinnitus-related decreases in postsynaptic gamma-amino butyric acid (GABA) signaling suggestive of GABA-A receptors (GABA-ARs) subunit switches or loss of GABA-ARs. VIP neurons favoring excitation of layer 5 PNs via disinhibition, were depolarized with significantly lower current to evoke action potentials (rheobase current). The excitability of VIP neurons was significantly increased, with this increase being strongly correlated to the rat’s tinnitus score. Tinnitus-related changes in nAChR signaling were then tested in layer 5 PNs and VIP neurons. Both PNs and VIP neurons receive cholinergic input from basal forebrain and were highly sensitive to nicotinic stimulation. Optical stimulation of basal forebrain (BF) terminals evoked a depolarizing current from VIP neurons. In tinnitus animals, layer 5 PNs showed a significant loss of nAChR signaling, while, VIP neurons showed tinnitus-related increase in responses to nicotinic stimulation. Most of the nAChR responses in auditory cortex are believed to be mediated via volume transmission of acetylcholine (ACh). Continuous voltage clamped recordings were used to examine the activity of excitatory and inhibitory neurons impacting PNs in the presence of bath applied ACh. We observed significant tinnitus-related changes in nAChR signaling with layer 5 PNs showing significantly larger GABAergic input after prolonged bath application of ACh. This led us to hypothesize that desensitization of nAChRs could increase/normalize the activity of GABAergic input neurons. To test this hypothesis, nAChR partial desensitizing agonists sazetidine-A and varenicline were used in cellular and behavioral studies. Immediately after bath application of sazetidine-A or varenicline, a dramatic increase in the activity of inhibitory input neurons onto PNs was observed. In a behavioral tinnitus test, both sazetidine-A and varenicline were effective in lowering the tinnitus-like behavior. In conclusion, we identified a significant tinnitus-related disruption in intrinsic physiology of layer 5 PNs and VIP neurons, with strong evidence of dysregulated cholinergic signaling. Partial/desensitizing agonists sazetidine-A and varenicline increased the activity of inhibitory input neurons, showing therapeutic potential in both subcellular and behavioral studies.

MicroRNAs (miRNAs) are short, non-coding RNA sequences that are generated from longer primary transcripts (pri-miRNA). These pri-miRNAs are processed by the endonuclease Drosha into a hairpin secondary structure (pre-miRNA), exported from the nucleus and cleaved by the enzyme Dicer to form a duplex RNA molecule. This miRNA:miRNA* duplex is subsequently further processed to form a single-stranded, mature miRNA. miRNAs have been extensively characterized and are known to play important roles in various physiologic and pathologic pathways. One hallmark of miRNAs function is their ability to modulate the downstream activities of protein-coding genes, as well as various other aspects of gene expression, by acting as post-transcriptional repressors of their messengerRNA (mRNA) targets. miR-184 is a highly conserved miRNA gene expressed in the Drosophila nervous system throughout development; and has been shown to target key regulators of differentiation, proliferation and apoptosis. Here we identify a novel role for miR-184 in regulating the development and maintenance of the Drosophila melanogaster post-embryonic nervous system. We present evidence which suggest miR-184 targets (i) paralytic (para), a voltage-gated sodium channel, shown to control neuronal excitability; and (ii) tramtrack69 (ttk69), a transcription factor known to regulate glial cell number and fate determination during embryonic development. In the absence of miR-184, homozygous loss-of-function mutant adult flies demonstrate hyperactive episodes in response to mechanical shock, indicative of increased susceptibility to seizures. Homozygous loss-of-function mutants also exhibit shortened lifespan, as well as reduced group longevity. Additionally, miR-184 deficient mutant larvae exhibit abnormal development of glia and glial progenitors; while expression of miR-184 exclusively in glia - reversed polarity- (repo) expressing cells - up-regulates development of glial cells. Phenotypes of the adult loss-of-function mutant are suppressed by genetic loss of para function; while larval phenotypes are rescued by reducing the genetic dosage of ttk69. These data imply that miR-184 functions to control post-embryonic gliogenesis, as well as in maintaining neuronal excitability and integrity of the Drosophila aging brain. / Biology

Opioids are used for the clinical treatment of pain, but can lead to tolerance and addiction. In this project we examined the role of the serotonin (5-HT) system originating from the dorsal raphe nucleus (DRN) during morphine exposure, withdrawal, abstinence and following an acute stressor capable of initiating behavioral relapse. Following four days of morphine exposure rats showed a preference for the morphine paired side of the conditioned place preference (CPP) chamber. After four days of morphine abstinence, rats showed no net preference for the morphine paired side. The next day rats were exposed to forced swim stress and returned to the CPP chamber where they demonstrated stress-induced reinstatement. Utilizing whole-cell patch-clamp we demonstrated an increase in the amplitude of inhibitory post-synaptic currents (IPSCs) in 5-HT DRN neurons, but not non 5-HT DRN neurons of morphine-conditioned subjects. Next the stress neurohormone corticotrophin releasing factor (CRF) was administered in vitro instead of forced swim. We found an increase in CRF-R2-mediated inward current of 5-HT DRN neurons in animals with a morphine history. From this experiment we concluded that morphine history sensitizes 5-HT DRN neurons to the GABAergic inhibitory effects of stress and to some of the effects of CRF. In the next series of experiments we surgically implanted either morphine or placebo pellets in rats for 72 hours to create physical dependence. The pellets were subsequently removed, and animals experienced up to seven days of abstinence with and without forced swim stress exposure. Real time quantitative PCR was used to measure the mRNA levels of genes at multiple points across this timeline. We examined genes involved in trophic support, stress responses and 5-HT regulation. We determined that mRNA levels for brain-derived neurotrophic factor (BDNF) and the BDNF receptor TrkB were downregulated after opiate exposure, and again following seven days of abstinence. Following seven days of abstinence there was a decrease in mRNA levels of the CRF-R1 receptor and an increase in mRNA levels of the CRF-R2 receptor. During acute opiate exposure there was a decrease in mRNA levels for the autoregulatory 5-HT1A receptor. Finally following forced swim, there was an increase in mRNA levels of the 5-HT synthesis enzyme TPH2. Collectively these results indicate that a morphine history in abstinent subjects may produce hypofunctioning of the 5-HT DRN system induced by multiple neurochemical mechanisms and this dysregulation may enhance vulnerability to stress-induced relapse. / Cell Biology