Pharmacological Dissection of Intrinsic Optical Signal Reveals a Functional Coupling between Synaptic Activity and Astrocytic Volume Transient

Abstract

The neuronal activity-dependent change in the manner in which light is absorbed or scattered in brain tissue is called the intrinsic optical signal (IOS), and provides label-free, minimally invasive, and high spatial (~100 µm) resolution imaging for visualizing neuronal activity patterns. IOS imaging in isolated brain slices measured at an infrared wavelength (>700 nm) has recently been attributed to the changes in light scattering and transmittance due to aquaporin-4 (AQP4)-dependent astrocytic swelling. The complexity of functional interactions between neurons and astrocytes, however, has prevented the elucidation of the series of molecular mechanisms leading to the generation of IOS. Here, we pharmacologically dissected the IOS in the acutely prepared brain slices of the stratum radiatum of the hippocampus, induced by 1 s/20 Hz electrical stimulation of Schaffer-collateral pathway with simultaneous measurement of the activity of the neuronal population by field potential recordings. We found that 55% of IOSs peak upon stimulation and originate from postsynaptic AMPA and NMDA receptors. The remaining originated from presynaptic action potentials and vesicle fusion. Mechanistically, the elevated extracellular glutamate and K + during synaptic transmission were taken up by astrocytes via a glutamate transporter and quinine-sensitive K2P channel, followed by an influx of water via AQP-4. We also found that the decay of IOS is mediated by the DCPIB- and NPPB-sensitive anion channels in astrocytes. Altogether, our results demonstrate that the functional coupling between synaptic activity and astrocytic transient volume change during excitatory synaptic transmission is the major source of IOS.

The molecular mechanism of synaptic activity‐induced astrocytic volume transient

“…In a supportive role, astrocytes maintain appropriate ionic concentrations in extracellular space, for example, by taking up potassium ions (K + ) during and after intense synaptic transmission (Simard & Nedergaard, 2004;Lambert et al 2008;Dallerac et al 2013). During this process, the water molecules initially follow the movement of K + via the astrocyte-specific water channel, aquaporin-4 (AQP4), resulting in a transient volume increase in the astrocyte (Simard & Nedergaard, 2004;Andrew et al 2007;Kitaura et al 2009;Nagelhus & Ottersen, 2013;Woo et al 2018Woo et al , 2019. Then the astrocytic volume returns to normal by extruding chloride ions (Cl − ) and osmolytes, possibly via anion channels (Abdullaev et al 2006;Mulligan & MacVicar, 2006;Okada et al 2009;Woo et al 2019), followed by a subsequent water efflux through AQP4.…”

“…Regarding the astrocytic K + uptake mechanism, many studies have revealed that the extracellular K + originates from the neuronal postsynaptic glutamate receptor channels that open and release during excitatory synaptic transmission (MacVicar & Hochman, 1991;Djukic et al 2007;Cheung et al 2015;Woo et al 2019). However, the exact molecular identity of the channel or transporter responsible for the K + uptake in astrocytes has not been determined.…”

“…Various potential molecular candidates have been proposed in the past; Kir4.1 (inwardly rectifying K + channel), NKCC1 (Na + , K + , 2Cl − cotransporter), Na + /K + -ATPase, and K2P (two-pore domain K + channels) (MacVicar et al 2002;Su et al 2002;Djukic et al 2007;Pasler et al 2007;Macaulay & Zeuthen, 2012). Likewise, the molecular identity of the anion channel which is responsible for the efflux of Cl − and osmolytes has not been determined, though it has been proposed that the volume regulated anion channel (VRAC) might be involved (Mulligan & MacVicar, 2006;Okada et al 2009;Han et al 2019;Woo et al 2019;Yang et al 2019).…”

Low-cost embedded system for optical imaging of intrinsic signals

“…Relative changes in reflected light intensity correspond to relative changes in the local concentrations of oxygenated hemoglobin and deoxygenated hemoglobin. OIS series of images represent a bidimensional functional map of hemodynamic signals, which are a proxy of neural activity [1]. Briefly, when a neuron fires, there is a localized consumption of oxygen revealed by an initial augmentation in deoxygenated hemoglobin.…”

“…Certainly, with larger animals, OIS needs to carry out OIS through the thinned skull with a rotative tool [3]. Can be extended to study optogenetically evoked signals, although it requires transgenic mice expressing channelrhodopsin (ChR2) [4], it can be performed in isolated brain slices ex vivo [1] or facilitated through the use of an intracranial transparent window [5]. OIS has been employed as a diagnostic tool, taking advantage of the intrinsic contrast provided by hemoglobin, which allows imaging without the addition of external contrast reagents.…”