Arianna Maffei - US grants

DESCRIPTION (provided by applicant): As children we need to learn an infinite set of skills. Our brain, the site where all information converges and is processed, shapes and is shaped by our perception. Synapses transfer environmental inputs to neurons and the integration of these signals determines the neuronal response to a specific stimulus (Huberman et al., 2006;Mrsic-Flogel et al., 2006). Neural plasticity modulates the strength of synapses when they are activated by relevant inputs. Plasticity is particularly important during brain development when periods of intense refinement of neural circuits - critical periods - lead to a mature and healthy functioning brain (Hubel and Wiesel, 1970;Katz and Shatz, 1996;Morales et al., 2002;Hensch, 2005). In primary visual cortex, one of the best studied models for experience-dependent plasticity, synapse specific forms of plasticity such as long term potentiation (LTP) or depression (LTD) are fundamental mechanisms for visual neuron function (Smith et al., 2009). The central role of inhibitory synaptic transmission in cortical circuits is demonstrated by the involvement of inhibitory synapses in the modulation of the onset and duration of critical periods (Hensch et al., 1998;Fagiolini and Hensch, 2000;Fagiolini et al., 2004). Inhibitory synaptic transmission mediated by fast spiking interneurons (FS) is directly affected by experience (Maffei et al., 2004;Maffei et al., 2006). A novel form of plasticity - LTP of FS to pyramidal neuron synapses, LTPi - is induced in layer 4 by changes in visual drive. LTPi might contribute to shaping visual cortical neurons responsiveness and receptive fields;but if induced at maximal, or saturating, levels, such as those measured after visual deprivation, it might promote loss of function (Maffei et al., 2006). LTPi induction properties are the subject of the present proposal. State of the art electrophysiological techniques will be used to dissect the mechanisms for induction of LTPi. More specifically we will determine 1) the dependence of LTPi induction on the timing of FS and pyramidal neuron activity;2) LTPi requirement for coincidence detection mechanisms;3) its dependence on the range of postsynaptic neuron activity - subthreshold vs. suprathreshold;4) LTPi dependence on presynaptic frequency of firing. We will also begin to investigate the cellular mechanisms for LTPi: the dependence on calcium influx and the intracellular pathways involved in GABAA receptor trafficking at synapses- PKA (Poisbeau et al., 1999) - and in the regulation of GABAA receptor function - PKC (Poisbeau et al., 1999;Brandon et al., 2000;Song and Messing, 2005). Quadruple concurrent patch clamp recordings will be obtained from pyramidal and FS to measure the strength, dynamics and plasticity of monosynaptic inhibitory connections. Specific drugs will be applied to prevent the induction of LTPi and to begin the identification of the cellular pathways involved in its induction. PUBLIC HEALTH RELEVANCE: Visual experience shapes cortical circuits during development. Synapses translate environmental inputs into signals that are integrated by neurons, which, in turn, produce appropriate responses to the sensory stimulus (Huberman et al., 2006;Mrsic-Flogel et al., 2006). The central role played by inhibitory synaptic transmission in visual cortical circuits is demonstrated by recent findings regarding inhibitory synapses and the modulation of the onset and duration of critical periods for plasticity (Hensch et al., 1998;Hensch, 2005). The maturation and plasticity of inhibitory synapses in visual cortex are essential for the processing of visual inputs and the preservation of balanced circuit excitability (Rozas et al., 2001;Morales et al., 2002;Hensch and Fagiolini, 2005).

DESCRIPTION (provided by applicant): The gustatory cortex (GC) receives modulatory inputs from multiple regions of the limbic system. Lateral hypothalamus, medial prefrontal cortex, mediodorsal thalamus and basolateral amygdala (BLA) are all known to send projections to GC (Saper 1982; Allen, Saper et al. 1991; Maffei, Haley et al. 2012). Among these inputs, those from the BLA are the ones whose functional significance has been studied the most. Pharmacological and electrophysiological experiments in alert animals have suggested that inputs from BLA are necessary for GC to process information pertaining to the hedonic value of gustatory stimuli (Piette, Baez- Santiago et al. 2012) and the anticipatory value of taste-predictive cues (Samuelsen, Gardner et al. 2012). In addition, plastic changes of the BLA-GC connection have been associated with taste aversion learning (Guzman-Ramos and Bermudez-Rattoni 2012). Despite the abundance of studies investigating the functional role of the BLA-GC connection (Jones, French et al. 1999; Grossman, Fontanini et al. 2008; Guzman-Ramos and Bermudez-Rattoni 2012; Piette, Baez- Santiago et al. 2012; Parkes and Balleine 2013), very little is known on the synaptic organization and plasticity of these inputs. Evidence from intracellular and extracellular recordings in vivo suggests that BLA-GC inputs might exert complex excitatory as well as inhibitory actions (Yamamoto, Azuma et al. 1984; Hanamori 2009; Stone, Maffei et al. 2011), yet no information is available on the synaptic mechanisms underlying these effects. Furthermore, while analysis of BLA evoked potentials in GC provides evidence for learning-related plasticity at this connection (Escobar, Chao et al. 1998; Jones, French et al. 1999; Escobar and Bermudez-Rattoni 2000; Rodriguez-Duran, Castillo et al. 2011), the mechanisms, rules and postsynaptic targets of this plasticity are unknown. Until recently it has been impossible to selectively activate BLA afferents in vitro to finely dissect th GC circuits recruited by amygdalar inputs. The availability of optogenetic tools has however changed the situation (Zhang, Gradinaru et al. 2010; Stuber, Sparta et al. 2011; Yizhar, Fenno et al. 2011; Britt and Bonci 2013; Wang, Kloc et al. 2013), finally allowing us the fundamental questions pertaining to the synaptic organization of amygdalar afferents to GC to be addressed. The experiments proposed here rely on these novel techniques, combined with in vitro whole cell patch clamp recordings, to directly measure the properties of amygdalar synapses onto pyramidal cells and inhibitory interneurons in GC. This methodological approach will be complemented with pharmacological and behavioral manipulations to test the following hypotheses: 1) BLA afferents make direct functional synapses onto different cell types within GC local circuits; 2) Synaptic inputs onto pyramidal neurons and inhibitory interneurons show activity-dependent plasticity; 3) The strength of BLA- GC synapses is affected by hedonic learning. Altogether these experiments will allow us to investigate the synaptic organization of BLA-GC inputs, their plasticity and the changes associated with aversion learning. This framework represents an entirely novel approach to the study of GC inputs in brain slices and promises to provide the first circuit-level description of amygdalar synapses in the gustatory cortex.

? DESCRIPTION (provided by applicant): How is gustatory information represented in the cortex? The answer to this question has been one of the mostly intensely debated topics in the field of gustatory neuroscience1-3. Two theories have been proposed and discussed over the past decades. According to the labeled-line theory the physiochemical identity of a gustatory stimulus is encoded by specific subsets of neurons, each of which is narrowly tuned to a single taste quality (i.e. salty, sweet, sour, bitter or umami)4-7. An alternative theory, the across-neuron pattern theory, postulates that taste coding relies on the combined activity of large ensembles of cortical neurons8. According to this theory, neurons do not need to be selective for specific taste qualities, instead each neuron can densely represent information by encoding multiple qualities3,9. Both theories are supported by experimental evidence. While a recent 2-photon calcium imaging study suggested the exclusive presence of narrowly tuned neurons in GC of anesthetized mice7; years of electrophysiological recordings in anesthetized and alert rodents demonstrate the presence of both narrowly tuned and densely coding neurons in GC10-13. Despite evidence that neurons using different coding strategies exist in GC, the debate is still polarized and no unifying view of taste coding in the cortex has emerged. The overarching goal of this proposal is to test the hypothesis that narrowly tuned and densely coding neurons reflect different stages of cortical processing. Specifically, we propose that the coding scheme varies depending on the cortical layers, with superficial layers (i.e., layers 2/3) featuring narroly tuned neurons and deep layers (i.e., layers 4 and 5) containing densely coding neurons. In addition, the experiments in this proposal aim at providing a mechanistic explanation for these differences, linking coding properties with laminar-dependent variations in inhibitory drive. The experiments will rely on a combination of sophisticated electrophysiological approaches to directly relate coding properties, balance between excitation and inhibition, and properties of local microcircuits. The framework of this grant is firmly grounded in the literature on cortical coding of sensory information14-17, and supported by our preliminary results, showing layer-specific differences in taste response properties and local connectivity. This research will help the field go beyond a dated debate and will move the discussion on taste coding toward a more circuit-oriented perspective. If successful this research will provide, for the first time, a unifid view of taste coding in cortical circuits of alert rodents.

Summary The perception of taste is a complex process that includes the chemosensory detection of a stimulus as well as information regarding its hedonic value, whether a stimulus is pleasant or unpleasant. The gustatory cortex is thought to be a central component of the taste system, and is hypothesized to integrate both of chemosensory and affective dimensions of taste stimuli. However, to date, the neuronal and circuit mechanisms involved in this process remain unclear. In this proposal, we will fill a gap in the current understanding of circuit underpinning for taste by investigating the synaptic mechanisms underlying the integration of thalamocortical and amygdalocortical inputs to the primary gustatory cortex. The thalamocortical projection from the taste thalamus provides the gateway for chemosensory information to the gustatory cortex, while the basolateral nucleus of the amygdala has been identified as a major contributor of hedonic information regarding taste stimuli. Taking advantage of cutting-edge experimental approaches for circuit and synaptic analysis, and of a well-established learning paradigm, conditioned taste aversion, that preserves the chemosensory identity, but alters a stimulus affective dimension, we will determine how sensory and hedonic components of taste are integrated in the gustatory cortical circuit. The approach we propose will begin to bridge the gap between the extensive behavioral/ pharmacological and cellular/molecular work that has significantly advanced the field, but without information about underlying synaptic and circuit mechanisms remains difficult to reconcile in an overarching framework. In addition, our study has important implication for public health. Many neurological and psychiatric disorders are associated with altered sensory perception, or by anhedonia. Taste perception in human and rodents is characterized by sensory and affective dimensions, therefore it provides a unique model for understanding common mechanism of sensory processing, as well as for investigating the circuit underpinning for the integration of sensory stimuli with their hedonic value. Therefore, results from our current study have the potential of identifying novel targets for the development of therapeutic intervention.

PROJECT SUMMARY Cortical circuits generate dynamic patterns of activity. One of the great challenges of modern neuroscience is to determine the circuit architectures that generate such dynamics patterns, and understand their genesis and functional significance. Most research on brain dynamics focused on stable patterns of activity showing continuous transitions (e.g., oscillations). However, in recent years there has been an increased interest on transient dynamics, including the ones resulting from the sequential switching between metastable states. Extracellular recordings of cortical ensembles indicated that sequences of metastable states, characterized by correlated changes in activity can be detected across subpopulations of neurons. Metastable states have been associated with specific cognitive or sensory variables, suggesting an important role for brain function. Metastability was also observed in the absence of any behavior or stimulation ? suggesting that metastable states may be generated locally and may reflect intrinsic architectures of cortical circuits. Despite evidence for their functional significance, little is known about metastable dynamics in cortical circuits. Indeed, lack of a coordinated and systematic approach to study both temporal and spatial signatures of these patterns has limited progress in this area. This proposal aims at developing an integrated experimental-computational platform for detecting metastable dynamics in cortical ensembles, inferring the circuit organizational principles underlying them, and understanding how plasticity affects metastability. Our team is formed by six PIs with complementary expertise in the experimental and computational approaches necessary to successfully accomplish this program. We will focus on circuits in the superficial layers of the gustatory portion of the insular cortex, a well-established model for understanding metastability. Our long-term goal is to generalize our findings to the study of transient dynamics in other cortical areas and understand their relevance for sensory, motor and/or cognitive tasks. Successfully accomplishing the proposed research will allow us to identify universal principles of collective network dynamics underlying behavior and experience-dependent learning.

Project Summary As animals transition from relying on their mother?s milk to foraging and consuming foods, they experience a large variety of new tastants. These experiences regulate their taste preferences later in life through a process that likely relies on plasticity in neural circuits associated with taste and feeding. The gustatory cortex (GC) is involved in processing taste information. It is required for taste-motivated behaviors and for learning about the chemosensory and affective dimensions of gustatory stimuli. The effects of early experiences with tastants on the development of taste preferences or the maturation of GC circuits has not been investigated. Our previous work in a different sensory cortex demonstrated a central role for inhibitory neurons in experience-dependent plasticity and postnatal circuit refinement. In this proposal, we will take advantage of our previous work and set out to investigate how the maturation of inhibition in GC contributes to the expression of taste preferences. The first part of the proposal will determine how taste preferences mature over the course of postnatal development and assess the role of experience with tastants on the modulation of taste preferences. Our preliminary observations suggest that there is a sensitive period for the experience-dependent modulation of taste preferences that is restricted to the weeks between the time weaning and young adulthood. We will also assess the time course of maturation of inhibitory circuits in GC and their sensitivity to experience with tastants. Finally, we will assess the relationship between the maturation of inhibition and the development of taste preferences using a variety of approaches including enzymatic and chemogenetic manipulations of inhibitory circuits? maturation. These studies will determine the role of inhibitory circuit in GC in the behavioral expression of taste preferences. While mechanisms underlying learning about the value or physical cues associated with tastes has been investigated in adulthood, the mechanisms leading to such a refined circuit in postnatal development have not been described. The results of these studies will indicate the role of early food experiences in determining taste-based choices throughout life.