Mapping Transgene Insertion Sites Reveals the α-Cre Transgene Expression in Both Developing Retina and Olfactory Neurons

The Tg(Pax6-cre,GFP)2Pgr (α-Cre) mouse (MGI:3052661) is a commonly used Cre line thought to be retinal-specific. Using targeted locus amplification (TLA), we mapped the insertion site of the transgene, and defined primers useful to deduce zygosity. Further analyses revealed four tandem copies of the transgene. The insertion site mapped to clusters of vomeronasal and olfactory receptor genes. Using R26R and Ai14 Cre reporter mice, we confirmed retinal Cre activity, but also detected expression in olfactory neurons, implicating the influence of neighbouring regulatory elements. RT-PCR and the buried food pellet (BFP) test did not detect any effects of the transgene on flanking genes in the nasal mucosa and retina. Together, these data precisely map α-Cre, show that it does not affect surrounding loci, but reveal previously unanticipated transgene expression in olfactory neurons. The α-Cre mouse can be a valuable tool in both retinal and olfactory research.

Energizing effects of bupropion on effortful behaviors in mice under positive and negative test conditions: modulation of DARPP-32 phosphorylation patterns

Motivational symptoms such as anergia, fatigue, and reduced exertion of effort are seen in depressed people. To model this, nucleus accumbens (Nacb) dopamine (DA) depletions are used to induce a low-effort bias in rodents tested on effort-based decision-making. We evaluated the effect of the catecholamine uptake blocker bupropion on its own, and after administration of tetrabenazine (TBZ), which blocks vesicular storage, depletes DA, and induces depressive symptoms in humans. Male CD1 mice were tested on a 3-choice-T-maze task that assessed preference between a reinforcer involving voluntary physical activity (running wheel, RW) vs. sedentary activities (sweet food pellet intake or a neutral non-social odor). Mice also were tested on the forced swim test (FST), two anxiety-related measures (dark–light box (DL), and elevated plus maze (EPM)). Expression of phosphorylated DARPP-32 (Thr34 and Thr75) was evaluated by immunohistochemistry as a marker of DA-related signal transduction. Bupropion increased selection of RW activity on the T-maze. TBZ reduced time running, but increased time-consuming sucrose, indicating an induction of a low-effort bias, but not an effect on primary sucrose motivation. In the FST, bupropion reduced immobility, increasing swimming and climbing, and TBZ produced the opposite effects. Bupropion reversed the effects of TBZ on the T-maze and the FST, and also on pDARPP32-Thr34 expression in Nacb core. None of these manipulations affected anxiety-related parameters. Thus, bupropion improved active behaviors, which were negatively motivated in the FST, and active behaviors that were positively motivated in the T-maze task, which has implications for using catecholamine uptake inhibitors for treating anergia and fatigue-like symptoms.

Feeding of Aulacomya atra Under Different Organic Matter Sources (Autochthonous and Allochthonous) in a Chilean Patagonia Fjord Ecosystem

Aulacomya atra is an active suspension feeder, spatially dominant in the shallow-water hard-bottom benthic communities of the Chilean Patagonia fjords. In this region, the vertical flux of autochthonous organic matter (OM) reaching the benthos is augmented by allochthonous OM both from a terrestrial origin and from intensive salmon farming. This mixed pool of OM represents a potential source of food for a variety of benthic consumers, but to date little is known about the degree of utilization of these materials by filter feeders organisms. In this context, feeding experiments on A. atra in Puyuhuapi Fjord, Chilean Patagonia, were conducted during summer and winter 2018–2019. These experiments were designed to determine ingestion rates (IR) of A. atra fed with autochthonous (bacterial and microplanktonic community) and allochthonous (salmon food pellet) OM. Additionally, samples of A. atra tissues and suspended particulate organic matter (SPOM) were taken from the study area for stable isotope analysis. Data from laboratory experiments indicated that A. atra can feed on both autochthonous and allochthonous OM, but higher IR were detected in individuals fed with salmon food pellets. Because the IR is sensitive to food particle density rather than specific type of food, diet preferences of A. atra in feeding experiments could not be determined. Stable isotope analyses indicate that A. atra in natural environment preferentially exploits food with an isotopic signal corresponding to autochthonous OM, highlighting the primary role of phytoplankton carbon in their diet. Extensive utilization of terrestrially derived OM is therefore unlikely, although utilization of OM derived from salmon farming is not precluded because of the overlap in isotopic signal between food pellets and marine plankton.

Implicitly Learned Higher Order Associations Differentiates Recent and Remote Retrieval of Temporal Order Memory

Memory of an ordered sequence of distinct events requires encoding the temporal order as well as the intervals that separates these events. In this study, using order place association task where the animal learns to associate the location of the food pellet to the order of entry into the event arena, we probe the nature of temporal order memory in mice. In our task, individual trials, become distinct events, as the animal is trained to form unique association between entry order and a correct location. The inter-trial intervals (> 30 mins) are chosen deliberately to minimise the working memory contributions. We develop this paradigm initially using 4 order place associates and later extend it to 5 paired associates. Our results show that animals not only acquire these explicit (entry order to place) associations but also higher order associations that can only be inferred implicitly from the temporal order of these events. As an indicator of such higher order learning during the probe trail the mice exhibit predominantly prospective errors that declines proportionally with temporal distance. On the other hand, prior to acquiring the sequence the retrospective errors are dominant. Additionally, we also tested the nature of such acquisitions when temporal order CS is presented along with flavour as a compound stimulus comprising of order and flavour both simultaneously being paired with location. Results from these experiments indicate that the animal learns both order-place and flavour-place associations. Comparing with pure order place training, we find that the additional flavour in compound training did not interfere with the ability of the animals to acquire the order place associations. When tested remotely, pure order place associations could be retrieved only after a reminder training. Further higher order associations representing the temporal relationship between the events is markedly absent in the remote time.

Key kinematic features in early training predict performance of adult female mice in a single pellet reaching and grasping task

Detailed analyses of overly trained animal models have been long employed to decipher foundational features of skilled motor tasks and their underlying neurobiology. However, initial trial-and-error features that ultimately give rise to skilled, stereotypic movements, and the underlying neurobiological basis of flexibility in learning, to stereotypic movement in adult animals are still unclear. Knowledge obtained from addressing these questions is crucial to improve quality of life in patients affected by movement disorders. We sought to determine if known kinematic parameters of skilled movement in humans could predict learning of motor efficiency in mice during the single pellet reaching and grasping assay. Mice were food restricted to increase motivation to reach for a high reward food pellet. Their attempts to retrieve the pellet were recorded for 10 minutes a day for continuous 4 days. Individual successful and failed reaches for each mouse were manually tracked using Tracker Motion Analysis Software to extract time series data and kinematic features. We found the number of peaks and time to maximum velocity were strong predictors of individual variation in failure and success, respectively. Overall, our approach validates the use of select kinematic features to describe fine motor skill acquisition in mice and establishes peaks and time to maximum velocity as predictive measure of natural variation in motion efficiency in mice. This manually curated dataset, and kinematic parameters would be useful in comparing with pose estimation generated from deep learning approaches.

Cue-motivated reward seeking is negatively regulated by expected reward magnitude in Pavlovian-instrumental transfer

The Pavlovian-instrumental transfer (PIT) paradigm is widely used to assay the motivational influence of reward-paired cues, which is reflected by their ability to stimulate instrumental reward-seeking behavior. Leading models of incentive learning assume that motivational value is assigned to cues based on the total amount of reward they signal (i.e., their state value). Based on recent findings, we lay out the alternative hypothesis that cue-elicited reward predictions may actually suppress the motivation to seek out new rewards through instrumental behavior in order to facilitate efficient retrieval of a reward that is already expected, before it is lost or stolen. According to this view, cue-motivated reward seeking should be inversely related to the magnitude of an expected reward, since there is more to lose by failing to secure a large reward than a small reward. We investigated the influence of expected reward magnitude on PIT expression. Hungry rats were initially trained to lever press for food pellets before undergoing Pavlovian conditioning, in which two distinct auditory cues signaled food pellet delivery at cue offset. Reward magnitude was varied across cues and groups. While all groups had at least one cue that signaled three food pellets, the alternate cue signaled either one (Group 1/3), three (Group 3/3), or nine food pellets (Group 3/9). PIT testing revealed that the motivational influence of reward-predictive cues on lever pressing varied inversely with expected reward magnitude, with the 1-pellet cue augmenting performance and the 3- and 9-pellet cues suppressing performance, particularly near the expected time of reward delivery. This pattern was mirrored by opposing changes in the food-port entry behavior, which varied positively with expected reward magnitude. We discuss how these findings may relate to cognitive control over cue-motivated behavior.

Muscarinic M1, but not M4, receptor antagonism impairs divided attention in male rats

Divided attention may be more important than ever to comprehend, given ubiquitous distractors in modern living. In humans, concern has been expressed about the negative impact of distraction in education, the home, and the workplace. While acetylcholine supports divided attention, in part via muscarinic receptors, little is known about the specific muscarinic subtypes that may contribute. We designed a novel, high-response rate test of auditory sustained attention, in which rats complete variable-ratio runs on one of two levers, rather than emitting a single response. By doing this, we can present a secondary visual distractor task during some trials, for which a correct nosepoke response is reinforced with a more palatable food pellet. The nonspecific muscarinic antagonist scopolamine impaired performance, and slowed and reduced lever press activity. We then explored antagonists that preferentially block the M1 and M4 subtypes, because these receptors are potential therapeutic targets for cognitive enhancers. Telenzepine, an M1-preferring antagonist, impaired divided attention performance, but not performance of the attention task without distraction. Telenzepine also had fewer nonspecific effects than scopolamine. In contrast, the M4-preferring antagonist tropicamide had no effects. Analysis of overall behavior also indicated that accuracy in the main attention task decreased as a function of engagement with the distractor task. These results implicate the M1 receptor in divided attention.

Learning a non-neutral conditioned stimulus: place preference in the crab Neohelice granulata

In the wild, being able to recognize and remember specific locations related to food sources and the associated attributes of landmarks is a cognitive trait important for survival. In the present work we show that the crab Neohelice granulata can be trained to associate a specific environment with an appetitive reward in a conditioned place preference task. After a single training trial, when the crabs were presented with a food pellet in the target quadrant of the training arena, they were able to form a long-term memory related to the event. This memory was evident at least 24 h after training and was protein-synthesis dependent. Importantly, the target area of the arena proved to be a non-neutral environment, given that animals initially avoided the target quadrant. In the present work we introduce for the first time an associative one-trial memory paradigm including a conditioned stimulus with a clear valence performed in a crustacean.

Food and water deprivation modulating function on activation and direction of behavior in Wistar rats

The effects of food or water deprivation on the consumption of those commodities has been extensively reported in the literature. The effect of the interaction of those deprivation conditions on food and water consumption and on the temporal organization and dynamics of behavior is less known. In this study, we evaluated the effects of different conditions of food and water deprivation on the spatio-temporal dynamics of behavior when food and water are concurrently available. Six rats were exposed to four different conditions: a) food deprivation, b) water deprivation, c) food and water deprivation and d) no deprivation. Experimental sessions consisted of simultaneously presenting a food pellet and a drop of water using a Concurrent Fixed Time 30 s schedule on two dispensers located on opposite walls of an extended experimental chamber. Local (number of drops of water and pellets consumed, head entries to dispensers) and translational (location, displacement routes) patterns were recorded. We found differential effects of the deprivation conditions on the aforementioned measures with no equivalent behavioral dynamics under food and water deprivation. The results are discussed in terms of the modulating function of deprivation conditions on measures of vigor and direction of behavior.activation; behavior dynamics; directionality, food deprivation; motivation; water deprivation.

Social isolation enhances cued-reinstatement of sucrose and nicotine seeking, but this is reversed by a return to social housing

Physical or perceived (i.e. loneliness) social isolation is increasing in Western cultures. Unfortunately, social isolation is associated with a range of negative physical and mental health outcomes, including increased incidence of obesity and smoking. Here we monitored the impact of social isolation on a range of physical measures, and then tested whether social isolation in adult rats changes how reward-related stimuli motivate sucrose- or nicotine-seeking. Socially isolated rats showed elevated baseline CORT, gained significantly less weight across the study, were more active in response to a novel or familiar environment. Isolated rats also acquired nose-poking for a food pellet more rapidly, and showed increased susceptibility to cue-, but not reward-induced reinstatement. Notably, these effects are partially mitigated by a return to group housing, suggesting that they are not necessarily permanent, and that a return to a social setting can quickly reverse any deficits or changes associated with social isolation. This study advances our understanding of altered reward-processing in socially isolated individuals and reiterates the importance of socialisation in the treatment of disorders such as overeating and addiction.