Aude Marzo

The Presence of Background Dopamine Signal Converts Long-Term Synaptic Depression to Potentiation in Rat Prefrontal Cortex

Executive functions of the brain are believed to require tonic dopamine inputs to the prefrontal cortex (PFC). It is unclear, however, how this background dopamine activity controls synaptic plasticity in the PFC, a possible underlying mechanism of executive functions. Using PFC slices, we show that pairing of dopamine with weak tetanic stimulation, a maneuver that otherwise induces NMDA receptor-independent long-term depression (LTD), induces long-term potentiation (LTP) when “primed” with dopamine. This “priming” occurs through the combined activation of D1 and D2 receptors and requires 12–40 min to develop. Moreover, concurrent synaptic activation of NMDA receptors during priming is necessary for this novel form of LTP. We suggest that a role of background dopamine signals in the PFC is to prevent high-frequency synaptic inputs from abnormally inducing LTD and to secure the induction of LTP.

Background Dopamine Concentration Dependently Facilitates Long-term Potentiation in Rat Prefrontal Cortex through Postsynaptic Activation of Extracellular Signal-Regulated Kinases

Altered levels of tonic/background dopamine in prefrontal cortex (PFC) may underlie modifications of executive cognitive function. We showed previously in rat PFC slices that exogenously supplied background dopamine facilitates induction of long-term potentiation (LTP), a possible cellular substrate for the long-term component of executive cognitive function. In the present study, we characterized cellular and molecular mechanisms underlying this modulatory dopamine effect. We show first that the LTP-facilitating effect of tonic/background dopamine follows an inverted-U shape concentration curve and that the effective level of background dopamine slowly activates postsynaptic extracellular signal-regulated kinases (ERKs) to facilitate LTP. Furthermore, we show the evidence that LTP-inducing high-frequency stimulation evokes endogenous release of dopamine in PFC slices. This fast dopamine serves as a trigger for LTP in the presence of the background dopamine. In its absence, the endogenous dopamine triggered, instead, long-term depression. These results indicate that appropriate levels of tonic/background dopamine serve to activate critical molecular factors in PFC neurons and thereby facilitate induction of synaptic potentiation.

Neuroplasticity Regulation by Noradrenaline in Mammalian Brain

The neuromodulator noradrenaline (NA) is released in almost all brain areas in a highly diffused manner. Its action is slow, as it acts through G protein-coupled receptors, but its wide release in the brain makes NA a crucial regulator for various fundamental brain functions such as arousal, attention and memory processes [102]. To understand how NA acts in the brain to promote such diverse actions, it is necessary to dissect the cellular actions of NA at the level of single neurons as well as at the level of neuronal networks. In the present article, we will provide a compact review of the main literatures concerning the NA actions on neuroplasticity processes. Depending on which subtype of adrenoceptor is activated, NA differently affects intrinsic membrane properties of postsynaptic neurons and synaptic plasticity. For example, β-adrenoceptor activation is mainly related to the potentiation of synaptic responses and learning and memory processes. α2-adrenoceptor activation may contribute to a high-order information processing such as executive function, but currently the direction of synaptic plasticity modification by α2-adrenoceptors has not been clearly determined. The activation of α1-adrenoceptors appears to mainly induce synaptic depression in the brain. But its physiological roles are still unclear: while its activation has been described as beneficial for cognitive functions, it may also exert detrimental effects in some brain structures such as the prefrontal cortex.

Activity-dependent spine morphogenesis: a role for the actin-capping protein Eps8.

Neuronal activity regulates the formation and morphology of dendritic spines through changes in the actin cytoskeleton. However, the molecular mechanisms that regulate this process remain poorly understood. Here we report that Eps8, an actin-capping protein, is required for spine morphogenesis. In rat hippocampal neurons, gain- and loss-of-function studies demonstrate that Eps8 promotes the formation of dendritic spines but inhibits filopodium formation. Loss of function of Eps8 increases actin polymerization and induces fast actin turnover within dendritic spines, as revealed by free-barbed end and FRAP assays, consistent with a role for Eps8 as an actin-capping protein. Interestingly, Eps8 regulates the balance between excitatory synapses on spines and on the dendritic shaft, without affecting the total number of synapses or basal synaptic transmission. Importantly, Eps8 loss of function impairs the structural and functional plasticity of synapses induced by long-term potentiation. These findings demonstrate a novel role for Eps8 in spine formation and in activity-mediated synaptic plasticity.

CELLULAR MECHANISMS OF LONG-TERM DEPRESSION INDUCED BY NORADRENALINE IN RAT PREFRONTAL NEURONS

Noradrenaline (NA) is released in the prefrontal cortex (PFC) during salient behavioral phases and thought to modulate PFC-mediated cognitive functions. However, cellular actions of NA in PFC neurons are still not well understood. In the present study, we investigated long-term effects of bath-applied NA (12.5 min) on glutamatergic synaptic transmission in rat PFC pyramidal neurons maintained in vitro. We found that NA concentration-dependently (5 microM< or =[NA]< or =20 microM) induces long-term depression (LTD) of layer I-II to layer V pyramidal neuron glutamatergic synapses. NA acts through alpha1- and alpha2-adrenoceptors, but not beta-adrenoceptors, to induce LTD. This NA-induced LTD depends on concurrent single synaptic activations of N-methyl-d-aspartate (NMDA) receptors and requires the activation of protein kinase C and postsynaptic Extracellular signal-Regulated Kinases (ERK1/2). Western blot analyses showed that NA (20 microM for 12.5 min) indeed induces transient increases of ERK1/2 phosphorylation in PFC neurons, which is dependent at least in part on the activation of NMDA receptors and alpha1-adrenoceptors. Together, these results demonstrate that NA can lastingly depress glutamatergic synapses in rat PFC neurons through mechanisms involving alpha-adrenoceptors, NMDA receptors, and the activation of postsynaptic ERK1/2.

Unilateral electrical stimulation of rat locus coeruleus elicits bilateral response of norepinephrine neurons and sustained activation of medial prefrontal cortex.

The brain stem nucleus locus coeruleus (LC) is thought to modulate cortical excitability by norepinephrine (NE) release in LC forebrain targets. The effects of LC burst discharge, typically evoked by a strong excitatory input, on cortical ongoing activity are poorly understood. To address this question, we combined direct electrical stimulation of LC (LC-DES) with extracellular recording in LC and medial prefrontal cortex (mPFC), an important cortical target of LC. LC-DES consisting of single pulses (0.1-0.5 ms, 0.01-0.05 mA) or pulse trains (20-50 Hz, 50-200 ms) evoked short-latency excitatory and inhibitory LC responses bilaterally as well as a delayed rebound excitation occurring ∼100 ms after stimulation offset. The pulse trains, but not single pulses, reliably elicited mPFC activity change, which was proportional to the stimulation strength. The firing rate of ∼50% of mPFC units was significantly modulated by the strongest LC-DES. Responses of mPFC putative pyramidal neurons included fast (∼100 ms), transient (∼100-200 ms) inhibition (10% of units) or excitation (13%) and delayed (∼500 ms), sustained (∼1 s) excitation (26%). The sustained spiking resembled NE-dependent mPFC activity during the delay period of working memory tasks. Concurrently, the low-frequency (0.1-8 Hz) power of the local field potential (LFP) decreased and high-frequency (>20 Hz) power increased. Overall, the DES-induced LC firing pattern resembled the naturalistic biphasic response of LC-NE neurons to alerting stimuli and was associated with a shift in cortical state that may optimize processing of behaviorally relevant events.

Wnt signalling tunes neurotransmitter release by directly targeting Synaptotagmin-1

The functional assembly of the synaptic release machinery is well understood; however, how signalling factors modulate this process remains unknown. Recent studies suggest that Wnts play a role in presynaptic function. To examine the mechanisms involved, we investigated the interaction of release machinery proteins with Dishevelled-1 (Dvl1), a scaffold protein that determines the cellular locale of Wnt action. Here we show that Dvl1 directly interacts with Synaptotagmin-1 (Syt-1) and indirectly with the SNARE proteins SNAP25 and Syntaxin (Stx-1). Importantly, the interaction of Dvl1 with Syt-1, which is regulated by Wnts, modulates neurotransmitter release. Moreover, presynaptic terminals from Wnt signalling-deficient mice exhibit reduced release probability and are unable to sustain high-frequency release. Consistently, the readily releasable pool size and formation of SNARE complexes are reduced. Our studies demonstrate that Wnt signalling tunes neurotransmitter release and identify Syt-1 as a target for modulation by secreted signalling proteins.

Evaluation of Synapse Density in Hippocampal Rodent Brain Slices

In the brain, synapses are specialized junctions between neurons, determining the strength and spread of neuronal signaling. The number of synapses is tightly regulated during development and neuronal maturation. Importantly, deficits in synapse number can lead to cognitive dysfunction. Therefore, the evaluation of synapse number is an integral part of neurobiology. However, as synapses are small and highly compact in the intact brain, the assessment of absolute number is challenging. This protocol describes a method to easily identify and evaluate synapses in hippocampal rodent slices using immunofluorescence microscopy. It includes a three-step procedure to evaluate synapses in high-quality confocal microscopy images by analyzing the co-localization of pre- and postsynaptic proteins in hippocampal slices. It also explains how the analysis is performed and gives representative examples from both excitatory and inhibitory synapses. This protocol provides a solid foundation for the analysis of synapses and can be applied to any research investigating the structure and function of the brain.