David Rosenegger

The perception of stress alters adaptive behaviours in Lymnaea stagnalis.

SUMMARY Stress can alter adaptive behaviours, and as well either enhance or diminish learning, memory formation and/or memory recall. We show here that two different stressors have the ability to alter such behaviours in our model system, Lymnaea stagnalis. One, a naturally occurring stressor– the scent of a predator (crayfish) – and the other an artificially controlled one – 25 mmoll–1 KCl – significantly alter adaptive behaviours. Both the KCl stressor and predator detection enhance long-term memory (LTM) formation; additionally predator detection alters vigilance behaviours. The predator-induced changes in behaviour are also accompanied by specific and significant alterations in the electrophysiological properties of RPeD1 – a key neuron in mediating both vigilance behaviours and memory formation. Naive lab-bred snails exposed to crayfish effluent (CE; i.e. the scent of the predator) prior to recording from RPeD1 demonstrated both a significantly reduced spontaneous firing rate and fewer bouts of bursting activity compared with non-exposed snails. Importantly, in the CE experiments we used laboratory-reared snails that have not been exposed to a naturally occurring predator for over 250 generations. These data open a new avenue of research, which may allow a direct investigation from the behavioral to the neuronal level as to how relevant stressful stimuli alter adaptive behaviours, including memory formation and recall.

Ecologically relevant stressors modify long-term memory formation in a model system

Stress can alter adaptive behaviours, and as well either enhance or diminish learning, memory formation and/or memory recall. We focus attention on how environmentally relevant stressors (e.g. predator detection, crowding, and low concentrations of environmental Ca(++)) alter memory formation in the pond snail, Lymnaea stagnalis. We specifically look at operant conditioning of aerial respiration and whether or not long-term memory forms following the acquisition of the learned event, not performing aerial respiration. We will also examine the strain differences in Lymnaea which allow or cause isolated populations to possess different heritable cognitive capabilities, as manifested by differing abilities to form long-term memory.

Canadian association of neurosciences review : Learning at a snail's pace

While learning and memory are related, they are distinct processes each with different forms of expression and underlying molecular mechanisms. An invertebrate model system, Lymnaea stagnalis, is used to study memory formation of a non-declarative memory. We have done so because: (1) We have discovered the neural circuit that mediates an interesting and tractable behaviour; (2) This behaviour can be operantly conditioned and intermediate-term and long-term memory can be demonstrated; and (3) It is possible to demonstrate that a single neuron in the model system is a necessary site of memory formation. This article reviews how Lymnaea has been used in the study of behavioural and molecular mechanisms underlying consolidation, reconsolidation, extinction and forgetting.

Enhancing memory formation by altering protein phosphorylation balance

In Lymnaea, aerial respiration can be operantly conditioned and depending on the training procedure employed two forms of memory can result: intermediate-term (ITM) and long-term memory (LTM). ITM, which persists for 3h, is dependent on de novo protein synthesis whilst LTM, which persists for at least 24 h, is dependent on both de novo protein synthesis and altered gene activity. A single 0.5 h training session (i.e. ITM-training) leaves behind a residual molecular memory trace, which a second bout of ITM-training can activate and boost it to a LTM. Here we extend this finding to show that either inhibiting protein phosphatase activity with okadaic acid (1 microM), or increasing protein kinase C (PKC) activity and therefore protein phosphorylation with bryostatin (0.25 ng/mL) treatment prior to ITM-training, results in a LTM. However, following right pedal dorsal 1 (RPeD1) soma ablation neither of these treatments are effective in producing LTM following ITM-training, indicating transcription is a necessity. These findings suggest that the balance between phosphorylation and dephosphorylation in neurons is a key factor for LTM formation.

A quantitative proteomic analysis of long-term memory

BackgroundMemory is the ability to store, retain, and later retrieve learned information. Long-term memory (LTM) formation requires: DNA transcription, RNA translation, and the trafficking of newly synthesized proteins. Several components of these processes have already been identified. However, due to the complexity of the memory formation process, there likely remain many yet to be identified proteins involved in memory formation and persistence.ResultsHere we use a quantitative proteomic method to identify novel memory-associated proteins in neural tissue taken from animals that were trained in vivo to form a long-term memory. We identified 8 proteins that were significantly up-regulated, and 13 that were significantly down-regulated in the LTM trained animals as compared to two different control groups. In addition we found 19 proteins unique to the trained animals, and 12 unique proteins found only in the control animals.ConclusionsThese results both confirm the involvement of previously identified memory proteins such as: protein kinase C (PKC), adenylate cyclase (AC), and proteins in the mitogen-activated protein kinase (MAPK) pathway. In addition these results provide novel protein candidates (e.g. UHRF1 binding protein) on which to base future studies.

A High Performance, Cost-Effective, Open-Source Microscope for Scanning Two-Photon Microscopy that Is Modular and Readily Adaptable

Two-photon laser scanning microscopy has revolutionized the ability to delineate cellular and physiological function in acutely isolated tissue and in vivo. However, there exist barriers for many laboratories to acquire two-photon microscopes. Additionally, if owned, typical systems are difficult to modify to rapidly evolving methodologies. A potential solution to these problems is to enable scientists to build their own high-performance and adaptable system by overcoming a resource insufficiency. Here we present a detailed hardware resource and protocol for building an upright, highly modular and adaptable two-photon laser scanning fluorescence microscope that can be used for in vitro or in vivo applications. The microscope is comprised of high-end componentry on a skeleton of off-the-shelf compatible opto-mechanical parts. The dedicated design enabled imaging depths close to 1 mm into mouse brain tissue and a signal-to-noise ratio that exceeded all commercial two-photon systems tested. In addition to a detailed parts list, instructions for assembly, testing and troubleshooting, our plan includes complete three dimensional computer models that greatly reduce the knowledge base required for the non-expert user. This open-source resource lowers barriers in order to equip more laboratories with high-performance two-photon imaging and to help progress our understanding of the cellular and physiological function of living systems.

The participation of NMDA receptors, PKC, and MAPK in the formation of memory following operant conditioning in Lymnaea

BackgroundMemory is the ability to store, retain, and later retrieve information that has been learned. Intermediate term memory (ITM) that persists for up to 3 h requires new protein synthesis. Long term memory (LTM) that persists for at least 24 h requires: DNA transcription, RNA translation, and the trafficking of newly synthesized proteins. It has been shown in a number of different model systems that NMDA receptors, protein kinase C (PKC) and mitogen activated protein kinase (MAPK) are all involved in the memory formation process.ResultsHere we show that snails trained in control conditions are capable of forming, depending on the training procedure used, either ITM or LTM. However, blockage of NMDA receptors (MK 801), inhibition of PKC (GF109203X hydrochloride) and MAPK activity (UO126) prevent the formation of both ITM and LTM.ConclusionsThe injection of either U0126 or GF109203X, which inhibit MAPK and PKC activity respectively, 1 hour prior to training results in the inhibition of both ITM and LTM formation. We further found that NMDA receptor activity was necessary in order for both ITM and LTM formation.

A slow or modulatory role of astrocytes in neurovascular coupling.

Astrocytes are thought to play an important role in NVC, a process that allows the brain to locally control blood flow in response to changes in activity. However, there is an ongoing debate as to when, and under what conditions astrocyte activity is required. In the following review we set forth the hypotheses that astrocytes: (i) act to modulate but not initiate functional hyperemia and (ii) help set the basal tone state of the brain microvasculature by the tonic release of vaso‐active messengers. Through these actions astrocytes could help match metabolic demand with supply over a spectrum of activity timescales.

Arteriole Dilation to Synaptic Activation that is Sub-Threshold to Astrocyte Endfoot Ca2+ Transients

Ca2+-dependent pathways in neurons and astrocyte endfeet initiate changes in arteriole diameter to regulate local brain blood flow. Whether there exists a threshold of synaptic activity in which arteriole diameter is controlled independent of astrocyte endfeet Ca2+ remains unclear. We used two-photon fluorescence microscopy to examine synaptically evoked synthetic or genetic Ca2+ indicator signals around penetrating arterioles in acute slices of the rat neocortex. We discovered a threshold below which vasodilation occurred in the absence of endfeet Ca2+ signals but with consistent neuronal Ca2+ transients, suggesting endfoot Ca2+ is not necessary for activity-dependent vasodilation under subtle degrees of brain activation.

Social transmission and buffering of synaptic changes after stress

Stress can trigger enduring changes in neural circuits and synapses. The behavioral and hormonal consequences of stress can also be transmitted to others, but whether this transmitted stress has similar effects on synapses is not known. We found that authentic stress and transmitted stress in mice primed paraventricular nucleus of the hypothalamus (PVN) corticotropin-releasing hormone (CRH) neurons, enabling the induction of metaplasticity at glutamate synapses. In female mice that were subjected to authentic stress, this metaplasticity was diminished following interactions with a naive partner. Transmission from the stressed subject to the naive partner required the activation of PVN CRH neurons in both subject and partner to drive and detect the release of a putative alarm pheromone from the stressed mouse. Finally, metaplasticity could be transmitted sequentially from the stressed subject to multiple partners. Our findings demonstrate that transmitted stress has the same lasting effects on glutamate synapses as authentic stress and reveal an unexpected role for PVN CRH neurons in transmitting distress signals among individuals.In mice, stress-induced priming of glutamate synapses in the PVN can be transmitted through social interactions. This requires PVN CRH neuron activation in both of the interacting mice and release of an alarm pheromone from the stressed mouse.