Abstract
Neurons exhibit strong coupling of electrochemical and metabolic activity. Increases in intrinsic fluorescence from either oxidized flavoproteins or reduced nicotinamide adenine dinucleotide (phosphate) [NAD(P)H] in the mitochondria have been used as an indicator of neuronal activity for the functional mapping of neural circuits. However, this technique has not been used to investigate the flow of olfactory information within the circuitry of the main olfactory bulb (MOB). We found that intrinsic flavoprotein fluorescence signals induced by electrical stimulation of single glomeruli displayed biphasic responses within both the glomerular (GL) and external plexiform layers (EPL) of the MOB. Pharmacological blockers of mitochondrial activity, voltage-gated Na+ channels, or ionotropic glutamate receptors abolished stimulus-dependent flavoprotein responses. Blockade of GABAA receptors enhanced the amplitude and spatiotemporal spread of the flavoprotein signals, indicating an important role for inhibitory neurotransmission in shaping the spread of neural activity in the MOB. Stimulus-dependent spread of fluorescence across the GL and EPL displayed a spatial distribution consistent with that of individual glomerular microcircuits mapped by neuroanatomic tract tracing. These findings demonstrated the feasibility of intrinsic fluorescence imaging in the olfactory systems and provided a new tool to examine the functional circuitry of the MOB.
Highlights
Functional optical imaging has been a valuable tool for examining the spatiotemporal connectivity of neuronal circuits
Vertical region of interests (ROIs) taken perpendicular to the main olfactory bulb (MOB) surface and centered on the stimulated glomerulus exhibited a stimulus-dependent signal that spread across the layers of the MOB over time (Fig 3A, right)
Horizontal ROIs taken from the middle of the GL and external plexiform layers (EPL) exhibited lateral stimulus-dependent signal spread over time (Fig 3A, bottom)
Summary
Animals and olfactory bulb slice preparationAll studies were approved by the University of Maryland, Baltimore IACUC committee. Animals (3–6 week old male and female C57BL6/J mice) were anesthetized with saturated isoflurane vapor and the olfactory bulbs surgically removed and immediately placed in 4 ̊C oxygenated sucrose-artificial cerebrospinal fluid (sucrose-ACSF) containing 26 mM NaHCO3, 1 mM NaH2PO4, 3 mM KCl, 5 mM MgSO4, 0.5 mM CaCl2, 10 mM glucose, and 248 mM sucrose, equilibrated with 95% O2-5% CO2, pH 7.38. Slices were incubated in oxygenated ACSF (containing 124 mM NaCl, 26 mM NaHCO3, 3 mM KCl, 1.25 mM NaH2PO4, 2 mM MgSO4, 2 mM CaCl2, and 15 mM glucose equilibrated with 95% O2-5% CO2, pH 7.4) at 30 ̊C for 20– 30 min at room temperature (22 ̊C) in ACSF for ~1 hr prior to use. The pharmacological agent, substrate change or washouts were perfused into the chamber for 15 min prior to the start of each recording. Pharmacological agents used for the experiments include mitochondrial toxin diphenyleneiodium (DPI), voltage-gated Na+ channel toxin tetrodotoxin (TTX), AMPA receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline-2,3-dione (NBQX), NMDA receptor antagonist (2R)-amino-
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
