Periodic Reporting for period 1 - NeuroSens (Neuromodulation of Sensory Processing) Reporting period: 2019-05-01 to 2021-07-31 Summary of the context and overall objectives of the project Cholinergic neuromodulation regulates arousal, learning and attention. Thereby it can modulate the way our brains interpret our sensory experiences. How our neural circuitry integrates and processes incoming sensory input to generate a meaningful output as well as how this is regulated by neuromodulation remains to be fully deciphered. This project aims to gain insights into neuromodulation of sensory processing by using the mouse primary somatosensory cortex (S1) as an experimental model. Mice use their whiskers for explorative behaviour and spatial navigation in a manner analogous to our use of fingers in somatosensory discrimination. Almost a third of the mouse neocortex is represented by S1 and a large portion of it processes sensory inputs from whiskers. The cellular mechanistic insights gained from the project will contribute to answering neurocomputational questions on the role and function of cortical microcircuit in sensory information processing. A greater understanding of sensory processing and its neuromodulation will also advance our understanding of neuropathology associated with several brain disorders that typify health burdens of today’s society. For example, deficits in sensory processing is found in many neurodevelopmental disorders, and drugs that are most effective in treating patients afflicted with Alzheimer’s are those that modulate actions of acetylcholine (ACh), the neuromodulator secreted by cholinergic neurons. The overall goal of the project is to understand cholinergic neuromodulation of sensory processing and the cellular mechanisms that govern it. Specifically, the neuromodulation of whisker-evoked responses in layer 2/3 pyramidal neurons of adult S1. To this end, its main objectives are 1) to characterise whisker-evoked responses in layer(L) 2/3 pyramidal neurons and 2) to examine the effects of ACh on these responses, and its mechanism. The whisker-evoked responses are cortical state dependent, and preliminary findings suggest that enhancement of cholinergic tone leads to a desynchronised cortical state. Current experiments underway involving pharmacological and optogenetic manipulation of cholinergic tone aim to further elucidate the cellular mechanisms that underlie this cortical desynchronisation and its effects on whisker-evoked responses. Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far State-of-the-art two photon guided in vivo whole cell recordings are carried out to characterise the whisker-evoked responses in L2/3 pyramidal neurons under urethane anaesthesia, which maintained a ‘controlled brain state’. Urethane anaesthesia mimics a slow wave sleep like brain state that is characterised by a cortical slow oscillation consisting of synchronous membrane potential fluctuations between a depolarised (‘up’) and hyperpolarised (‘down’) state. ACh strongly influences cortical network activity, and a low cholinergic tone is found during slow wave sleep.The whisker-evoked responses are elicited by application of an air puff to the whiskers, which resulted in their deflection (i.e. protraction and retraction of whiskers that approximate their natural whisking behaviour). The frequency and number of presented stimuli are varied, e.g. a single stimulus presented at varying frequency or a train of stimuli. Discharge of action potentials (APs) in L2/3 pyramidal neurons is sparse, and <10% of neurons fired APs in response to whisker puff(s) being delivered. The evoked responses in L2/3 pyramidal neurons that did not fire APs to whisker puff(s) are heterogeneous and cortical state dependent. If a cortical down state precedes the stimulus a subthreshold postsynaptic potential in L2/3 pyramidal neurons is evoked. Such subthreshold events are known as dendritic spikes, which are regenerative events produced by non-linear integration of synaptic inputs onto the dendritic arbour of a neuron and at times by backpropagating APs from the soma into dendrites. In contrast if a whisker puff is presented during a cortical upstate the membrane potential of L2/3 pyramidal neuron undergoes hyperpolarisation. The evoked responses to a train of whisker puffs are complex: from eliciting sharply delineated postsynaptic potentials corresponding to each individual whisker deflection to summating of responses with each subsequent whisker deflection. To localise the specific basal forebrain nucleus that sends cholinergic projections to S1, a combination of anterograde viral tracers and retrograde neuroanatomical tracers is carried out in a transgenic mouse line where site specific expression of cholinergic neurons can be driven. A relatively small proportion of cholinergic neurons from this basal forebrain nucleus sends a large network of diffused axonal projections to S1. The viral injection of channelrhodopsin2, an opsin that is light sensitive, into this nucleus results in the specific expression of channelrhodopsin2 in these cholinergic axons; thereby permitting manipulation of cholinergic tone in S1 using light (i.e. optogenetics) during whisker deflections to examine cholinergic modulation of whisker-evoked responses (characterised above). Initial findings with pharmacological manipulation of cholinergic tone during in vivo whole cell recordings show that activation of ACh receptors leads to depolarisation and increased rate of firing of L2/3 pyramidal neurons in response to somatic current injections. The application of an ACh receptor agonist also induces a highly desynchronised brain state altering the synchronous cortical up/down state transitions. State-of-the-art in vivo whole cell recording and calcium imaging experiments are currently underway to examine the effects of pharmacological and optogenetic manipulation of cholinergic tone on previously characterised whisker-evoked responses. The python scripted codes developed in collaboration with James Thomas at Jean Golding Institute, University of Bristol for the analyses of in vivo data generated from this project are currently maintained in a private GitHub repository, which will be made public at the initial publication of the project on the preprint server for biology, bioRxiv. The final publication will be made open access with a link to the GitHub repository included. Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far) The project aimed to examine processing of sensory information and their neuromodulation. The responses to sensory stimuli are dependent on cortical state and increasing cholinergic tone leads to cortical desynchronisation. It is considered that the desynchronisation of cortical activity through the engagement of cholinergic pathway enhances information processing via reducing redundancy during alert and attentive conditions. The current experiments in which the cholinergic tone is modulated via pharmacologically and/or optogenetically during sensory stimuli presentation are expected to yield results that will shed light on the cellular and circuit mechanisms involved in neuromodulation of sensory processing. These findings will contribute to European and international brain mapping initiatives to investigate, model and map the functional connectivity of the entire brain (e.g. Blue Brain Project, BRAIN Initiative). A greater mechanistic understanding of neuromodulation of sensory processing will also shift our knowledge frontier on neuropathology of several brain disorders (that are described above), which will ultimately lead to a positive impact socio-economically given the global burden of disease posed by these disorders.