Region-Specific Differences in Morphometric Features and Synaptic Colocalization of Astrocytes During Development

Abstract

It is well established that astrocytes play pivotal roles in neuronal synapse formation and maturation as well as in the modulation of synaptic transmission. Despite their general importance for brain function, relatively little is known about the maturation of astrocytes during normal postnatal development, especially during adolescence, and how that maturation may influence astroglial-synaptic contact. The medial prefrontal cortex (mPFC) and dorsal hippocampus (dHipp) are critical for executive function, memory, and their effective integration. Further, both regions undergo significant functional changes during adolescence and early adulthood that are believed to mediate these functions. However, it is unclear the extent to which astrocytes change during these late developmental periods, nor is it clear whether their association with functional synapses shifts as adolescent and young adult maturation proceeds. Here we utilize an astrocyte-specific viral labeling approach paired with high resolution single cell astrocyte imaging and threedimensional reconstruction to determine whether mPFC and dHipp astrocytes have temporally distinct maturation trajectories. mPFC astrocytes, in particular, continue to mature well into emerging adulthood (postnatal day 70). Moreover, this ongoing maturation is accompanied by a substantial increase in colocalization of astrocytes with the postsynaptic neuronal marker, PSD-95. Taken

High‐Resolution Three‐Dimensional Imaging of Individual Astrocytes Using Confocal Microscopy

“…Meanwhile, Lck‐GCaMP constructs have revealed the dynamics of astrocyte calcium signaling at the PAPs (Shigetomi et al., 2013; Shigetomi, Kracun, & Khakh, 2010). Recently, three‐dimensional reconstructions of Lck‐GFP‐transduced astrocytes were used to explore morphometric changes after administration of various drugs of abuse (Kruyer et al., 2019; Scofield et al., 2016; Siemsen et al., 2019; Testen et al., 2018) as well as for measurements of developmental changes (Testen et al., 2019).…”

“…Results are later plotted. See Figure 3 of Testen et al (2018) and Figures 4 and 5 of Testen et al (2019) for more information and examples of the types of data that one can obtain.…”

“…In order to more completely observe structural properties of astrocytes, our laboratory and others have utilized a lymphocyte‐specific protein tyrosine kinase (Lck) tagged with green fluorescent protein (GFP) (Scofield et al., 2016; Shigetomi, Kracun, Sofroniew, & Khakh, 2010; Testen et al., 2019; Testen, Sepulveda‐Orengo, Gaines, & Reissner, 2018). Post‐translational modification of the N‐terminus of Lck results in association with the plasma membrane.…”

“…In addition, immunohistochemistry (IHC) for synaptic markers such as synapsin‐1 and post‐synaptic density‐95 can be employed to assess synaptic colocalization of Lck‐GFP. Using these methods, our laboratory and others have shown changes in nucleus‐accumbens astrocytes associated with self‐administration of cocaine (Scofield et al., 2016; Testen et al., 2018), heroin (Kruyer, Scofield, Wood, Reissner, & Kalivas, 2019), and methamphetamine (Siemsen et al., 2019) as well as in other structures across development (Testen et al., 2019) and in response to severe stress (Jones, Paniccia, Lebonville, Reissner, & Lysle, 2018). The general methods employed in the above‐mentioned studies are described in the protocols below.…”

Molecular Plasticity of the Nucleus Accumbens Revisited—Astrocytic Waves Shall Rise

“…1), astrocytes are ideally positioned to control the plasticity of mixed Gluergic/GABAergic synapses. Depending on local activation and propagating Ca 2+ waves through the astrocytic syncytium due to high-frequency stimulation, cocaine exposure, lactation, or other stimuli, NAc astrocytes can adjust their morphology [155, 205, 206] resulting in different coverages of the synapse. Since Glu uptake and spillover is crucially dependent on astrocytic coverage, the tightness of astrocyte wrapping of the synapse can finely tune the balance between inhibitory and excitatory outcomes (Fig.…”