An updated suite of viral vectors for in vivo calcium imaging using intracerebral and retro-orbital injections in male mice

Abstract

Genetically encoded Ca2+ indicators (GECIs) are widely used to measure neural activity. Here, we explore the use of systemically administered PHP.eB AAVs for brain-wide expression of GECIs and compare the expression properties to intracerebrally injected AAVs in male mice. We show that systemic administration is a promising strategy for imaging neural activity. Next, we establish the use of EE-RR- (soma) and RPL10a (Ribo) soma-targeting peptides with the latest jGCaMP and show that EE-RR-tagged jGCaMP8 gives rise to strong expression but limited soma-targeting. In contrast, Ribo-tagged jGCaMP8 lacks neuropil signal, but the expression rate is reduced. To combat this, we modified the linker region of the Ribo-tag (RiboL1-). RiboL1-jGCaMP8 expresses faster than Ribo-jGCaMP8 but remains too dim for reliable use with systemic virus administration. However, intracerebral injections of the RiboL1-tagged jGCaMP8 constructs provide strong Ca2+ signals devoid of neuropil contamination, with remarkable labeling density.

High fidelity sensory-evoked responses in neocortex after intravenous injection of genetically encoded calcium sensors

“…However, to our knowledge no previous studies have demonstrated retinotopic and orientation-selective responses at the single-neuron level or the high-fidelity trial-to-trial responses achievable with intravenous injections of AAV-PhP.eB-Synapsin-GCaMP7s. Even the most recent soma-targeting AAV-PhP.eB-GCaMP8 study examined visual cortical responses to orientation and direction selective stimuli by intracranial injection of the AAV, not intravenous injection (Grodem et al, 2023).…”

“…In all past studies examining AAV-PhP.B- and AAV-PhP.eB-mediated GCaMP, the maximum post-injection period examined was 10 weeks (Allen et al, 2017; Hillier et al, 2017; Michelson et al, 2019; Brown et al, 2021; Grodem et al, 2023). We have extended this timeframe by demonstrating that expression remains stable and free of nuclear labeling for at least 34 weeks after transduction.…”

“…However, several new AAV variants cross the blood brain barrier in all mouse strains and even in non-rodent species, including non-human primates (Ravindra Kumar et al, 2020; Goertsen et al, 2022). It is expected that refinement of these and other newly developed tools, such as soma- and axon-targeted expression of GCaMP (Grodem et al, 2023), will make two-photon calcium imaging in all mammalian species completely ‘plug-and-play’ in the near future.…”

“…To change the transgene carried by an AAV capsid is straightforward, whereas creating new transgenic animal lines is a difficult and lengthy process. This flexibility is especially advantageous for calcium imaging because new generations of GECIs with enhanced performance and cell-type specific promoters are regularly engineered (Grodem et al, 2023). Additionally, intravenous AAV injections could be used to label neurons in other transgenic lines, e.g., disease models (as noted above) without the need for crossbreeding with a transgenic GECI line.…”

Advances in cellular resolution microscopy for brain imaging in rats

“…For example, neuropil contamination could be reduced by expression of soma restricted calcium sensors. 141,142 In addition, simultaneous imaging of multiple cell types could be achieved by restricting sensors to readily differentiable cellular compartments, such as axons and soma. Finally, imaging of apical dendrites could allow access to deep cortical neurons, an approach used to support population imaging in macaques.…”