Rapid stimulus-evoked astrocyte Ca ²⁺ elevations and hemodynamic responses in mouse somatosensory cortex in vivo

Abstract

Increased neuron and astrocyte activity triggers increased brain blood flow, but controversy exists over whether stimulationinduced changes in astrocyte activity are rapid and widespread enough to contribute to brain blood flow control. Here, we provide evidence for stimulus-evoked Ca 2+ elevations with rapid onset and short duration in a large proportion of cortical astrocytes in the adult mouse somatosensory cortex. Our improved detection of the fast Ca 2+ signals is due to a signal-enhancing analysis of the Ca 2+ activity. The rapid stimulation-evoked Ca 2+ increases identified in astrocyte somas, processes, and end-feet preceded local vasodilatation. Fast Ca 2+ responses in both neurons and astrocytes correlated with synaptic activity, but only the astrocytic responses correlated with the hemodynamic shifts. These data establish that a large proportion of cortical astrocytes have brief Ca 2+ responses with a rapid onset in vivo, fast enough to initiate hemodynamic responses or influence synaptic activity. and associated astrocytes, which causes fluctuations in cerebral blood flow (CBF) (1-5). Astrocytes are ideally situated for controlling activity-dependent increases in CBF because they closely associate with synapses and contact blood vessels with their end-feet (1, 6). Whether or not astrocytic Ca 2+ responses develop often or rapidly enough to account for vascular signals in vivo is still controversial (7-10). Ca 2+ responses are of interest because intracellular Ca 2+ is a key messenger in astrocytic communication and because enzymes that synthesize the vasoactive substances responsible for neurovascular coupling are Ca 2+ -dependent (1, 4). Neuronal activity releases glutamate at synapses and activates metabotropic glutamate receptors on astrocytes, and this activation can be monitored by imaging cytosolic Ca 2+ changes (11). Astrocytic Ca 2+ responses are often reported to evolve on a slow (seconds) time scale, which is too slow to account for activity-dependent increases in CBF (8,10,12,13). Furthermore, uncaging of Ca 2+ in astrocytes triggers vascular responses in brain slices through specific Ca 2+ -dependent pathways with a protracted time course (14, 15). More recently, stimulation of single presynaptic neurons in hippocampal slices was shown to evoke fast, brief, local Ca 2+ elevations in astrocytic processes that were essential for local synaptic functioning in the adult brain (16,17). This work prompted us to reexamine the characteristics of fast, brief astrocytic Ca 2+ signals in vivo with special regard to neurovascular coupling, i.e., the association between local increases in neural activity and the concomitant rise in local blood flow, which constitutes the physiological basis for functional neuroimaging.Here, we describe how a previously undescribed method of analysis enabled us to provide evidence for fast Ca 2+ responses in a main fraction of astrocytes in mouse whisker barrel cortical layers II/III in response to somatosensory stimulation. The astrocytic Ca 2+ responses were brief en...

Genetic targeting of astrocytes in the gliovascular unit of the adult brain

“…By using GECIs, Otsu and colleagues [6] recently observed that Ca 2+ responses in astrocyte processes precede hemodynamic activity in the olfactory bulb. This corroborates studies that demonstrated that astrocyte process Ca 2+ activity operates on a different timescale in comparison to the somata 34,40 .…”

“…However, using the same dye loading techniques coupled with in vivo Ca 2+ imaging, researchers were not able to confirm whether or not astrocyte Ca 2+ activity is indeed linked to hemodynamic responses 32,33 . To explain these conflicting studies, it is thought that Ca 2+ -mediated signaling in the distal, finer astrocytic processes are thought to be particularly important for neuro-glio-vascular signaling 34 . Importantly, one major caveat to previous dye-loading studies is that this technique is mostly effective for analyzing the astrocyte soma rather than its finer distal processes 35 .…”

IGF‐1 deficiency impairs neurovascular coupling in mice: implications for cerebromicrovascular aging

“…The signal was amplified with an AC/DC differential amplifier (high pass at 1 Hz, low pass at 1 kHz) (Model 3000; A‐M Systems, Inc. Carlsborg, WA, USA) and digitalized by the PowerLab/Labchart data acquisition system (ADInstruments, Colorado Springs, CO, USA) with a sampling rate of 40 kHz. Basal activity was analyzed as distribution of wave amplitude as a function of frequency (Park et al ., 2008), and the negative amplitude in the somatosensory evoked field potential response was considered as the excitatory postsynaptic potential (fEPSP) (Lind et al ., 2013). Analyses were performed on the average of 10 stimulation trials.…”

Pharmacologically induced impairment of neurovascular coupling responses alters gait coordination in mice

“…Analyses were performed on the average of 10 stimulation trials. The negative amplitude in the somatosensory-evoked field potential response was considered as the excitatory postsynaptic potential (fEPSP) (Lind et al 2013). Experiments lasted~20-30 min per mouse, which permitted stable physiological parameters to be obtained.…”