Dustin M. Graham - US grants

[unreadable] DESCRIPTION (provided by applicant): The suprachiasmatic nucleus (SCN) receives information about ambient light levels through the retinohypothalamic tract. This information is used to reset the molecular clock of individual SCN neurons, leading to entrainment of overt animal behavior and physiology. Recently, a new class of intrinsically photosensitive retinal ganglion cells (ipRGC's) utilizing melanopsin as their photopigment was discovered and found to provide the majority of retinal input to the SCN. However, many questions remain about how these melanopsin ganglion cells communicate with individual SCN neurons. One complicating factor is that classic rod and cone pathways also provide input to the SCN, relayed by melansopin ganglion cells and conventional ganglion cell types. Due to difficulties in recording in vivo light-evoked responses in the SCN, little is known about the organization and function of these various input pathways and how they lead to resetting of the biological clock. To circumvent this issue, we have developed a novel in vitro SCN brain- slice preparation that maintains functional connectivity to both retinas, enabling patch-clamp recordings from visually identified SCN neurons that are capable of responding to light. This allows us to conduct experiments concerning light processing in the SCN with unprecedented control and knowledge of the cells we record from. One of the main advantages is our ability to target cells for recording in the dorsal shell or ventrolateral core of the SCN, two sub-regions thought to play different roles in processing retinal input. Utilizing this novel slice preparation, the goal of the proposed study is to delineate the functional organization of rod/cone and melansopin based input to the SCN using patch-clamp recording techniques in combination with pharmacology, and to understand how these various inputs shape light responses from individual SCN cells. In addition, the function of endogenous PACAP, a neuropeptide found in melanopsin ganglion cells, will be characterized during processing of visual information in the SCN. Because of the similarities between rat and human circadian systems, studying how the SCN processes retinal input in our novel slice preparation will provide valuable information about the cellular basis of circadian rhythms, photic entrainment, and sleep regulation in humans. This will hopefully lead to standards of work conditions and schedules for millions of shift workers where disruption of circadian rhythms and sleep deprivation is a major occupational hazard, and in the case of hospitals, a threat to public health. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): The exact nature and mechanism of encoding chemical stimuli in primary gustatory cortex is a matter of debate. Chronic extracellular recordings from awake-behaving rats suggest a dynamic and distributed code 4-10. This model emphasizes the unique spatio/temporal patterns of activity among groups of broadly tuned neurons acting as dynamic ensembles. However, recent data from anesthetized mice using in-vivo calcium imaging show a precise gustatopic map of primary tastes in the cortex 1. This suggests a coding scheme of sharply tuned gustatory cortical neurons arranged in topographically organized clusters. While the evidence for an organized cortical map representing primary tastes is intriguing, enthusiasm for such a map is tempered by the reliance on anesthetized animals 1-3. Anesthesia is well known to cause aberrant and artificial cortical network dynamics, significantly affecting processing of sensory stimuli 11. This proposal will resolve this problem by directly testing the general hypothesis that taste responsive neurons in gustatory cortex from awake-behaving mice are narrowly tuned and topographically organized. This will be accomplished by using a unique combination of in vivo extracellular and whole-cell patch-clamp electrophysiology, with a novel head-restrained mouse preparation that allows precise and systematic targeting of recordings throughout gustatory cortex in awake-behaving mice. The specific aims will study the functional organization of the gustatory cortex in awake-behaving animals with unprecedented detail, from the network-level to detailed synaptic mechanisms. Aim 1: Taste responses in gustatory cortex are narrowly tuned and functionally organized into topographic clusters in awake-behaving mice. Aim 2: Single-unit taste responses reflect mapped multi-unit activity. Aim 3: A balance of excitatory and inhibitory synaptic conductances drive taste responses in gustatory cortical neurons and shapes tuning. PUBLIC HEALTH RELEVANCE: Diabetes is a widespread and costly public health problem in the US. A better understanding of the taste system and processing of sweet tasting foods will help us enact better, scientifically guided, policies for public nutrition, with the goal of reducig childhood and adult type-2 diabetes.