Retrograde optogenetic characterization of the pontospinal module of the locus coeruleus with a canine adenoviral vector

Abstract

Noradrenergic neurons of the brainstem extend projections throughout the neuraxis to modulate a wide range of processes including attention, arousal, autonomic control and sensory processing. A spinal projection from the locus coeruleus (LC) is thought to regulate nociceptive processing. To characterize and selectively manipulate the pontospinal noradrenergic neurons in rats, we implemented a retrograde targeting strategy using a canine adenoviral vector to express channelrhodopsin2 (CAV2-PRS-ChR2-mCherry). LC microinjection of CAV2-PRS-ChR2-mCherry produced selective, stable, transduction of noradrenergic neurons allowing reliable opto-activation in vitro. The ChR2-transduced LC neurons were opto-identifiable in vivo and functional control was demonstrated for >6 months by evoked sleep-wake transitions. Spinal injection of CAV2-PRS-ChR2-mCherry retrogradely transduced pontine noradrenergic neurons, predominantly in the LC but also in A5 and A7. A pontospinal LC (ps:LC) module was identifiable, with somata located more ventrally within the nucleus and with a discrete subset of projection targets. These ps:LC neurons had distinct electrophysiological properties with shorter action potentials and smaller afterhyperpolarizations compared to neurons located in the core of the LC. In vivo recordings of ps:LC neurons showed a lower spontaneous firing frequency than those in the core and they were all excited by noxious stimuli. Using this CAV2-based approach we have demonstrated the ability to retrogradely target, characterise and optogenetically manipulate a central noradrenergic circuit and show that the ps:LC module forms a discrete unit.This article is part of a Special Issue entitled SI: Noradrenergic System.

Dynamic Lateralization of Pupil Dilation Evoked by Locus Coeruleus Activation Results from Sympathetic, Not Parasympathetic, Contributions

“…We first examined if activation of the LC-NE neurons using electrical microstimulation would alter cortical arousal by shifting the power spectrum of cortical EEG, as had been previously shown using pharmacological compounds (Berridge and Foote, 1991; Steriade et al, 1993; Vazey and Aston-Jones, 2014) and photostimulation (Li et al, 2016). Consistent with previous work, brief electrical microstimulation (50 Hz, 6 biphasic current pulses, 200 µs per phase, 60 or 120 µA amplitude) of the LC induced the EEG power spectrum to shift towards higher frequencies (Figure 3A), quantifiable by an increase in the ratio of 10–100 Hz to 1–10 Hz power ( p < 0.02, Wilcoxon signed-rank test, Figure 3B).…”

The role of working memory and declarative memory in trace conditioning

“…For example, pathway specific manipulation may be achieved using two vectors by infusing a retrograde canine adenovirus-2 into a target region (e.g., mPFC) to deliver Cre-recombinase and an adeno-associated virus carrying a designer receptor into a target region that projects to the first region (e.g., VH) (Boender et al, 2014). Likewise, optogenetics also allows for similar pathway specific manipulation (Li et al, 2016; Walsh et al, 2012). The temporal resolution provided by optogenetics may be particularly useful given the defined CS-US onset-offset parameters of trace conditioning.…”

Deep Brain Stimulation Improves the Symptoms and Sensory Signs of Persistent Central Neuropathic Pain from Spinal Cord Injury: A Case Report

“…Recent optogenetic studies have shown that focal activation of the LC, whose major inputs are from the PFC, amygdala, PAG and dorsal noradrenergic bundle, may be either pro- or anti-nociceptive, suggestive of different neuron subgroups within the LC (Hickey et al, 2014). Consistent with this, recently Li et al (2016), used a canine adenoviral vector expressing channelrhodopsin2 to target ponto-spinal NE neurons, and found a subset of ventral LC neurons with spinal projections that is likely involved in the regulation of nociception. Interestingly, this subgroup of LC neurons had lower average firing rate than other noradrenergic neurons of the LC.…”

“…Most evidence available supports the PAG as a useful stimulation target (Pereira and Aziz, 2014). In addition to ascending somatosensory projections, the PAG receives significant inputs from the prefrontal cortex (PFC) and amygdala and then projects to nucleus raphe magnus (NRM) and locus coeruleus (LC) in the midbrain and dorsal horn neurons in the spinal cord (Van Bockstaele et al, 1991; Li et al, 2016). Recent case series on DBS for chronic neuropathic pain, which have utilized superior targeting methods compared to earlier studies and multi-day trials with an externalized electrode prior to final implantation, suggest clinically-relevant improvements of pain symptoms in 67%–83% of patients with heterogeneous types of pain (Boccard et al, 2013).…”

“…Norepinephrine (NE) has also been implicated in the modulation of pain (Hickey et al, 2014; Li et al, 2016). NE reuptake inhibitors or combinations of NE and 5-HT reuptake inhibitors are common therapies for CNP and are recommended first-line treatments for patients with SCI (Guy et al, 2016).…”