Optogenetic approaches to characterize the long-range synaptic pathways from the hypothalamus to brain stem autonomic nuclei

Abstract

Recent advances in optogenetic methods demonstrate the feasibility of selective photoactivation at the soma of neurons that express channelrhodopsin-2 (ChR2), but a comprehensive evaluation of different methods to selectively evoke transmitter release from distant synapses using optogenetic approaches is needed. Here we compared different lentiviral vectors, with sub-population-specific and strong promoters, and transgenic methods to express and photostimulate ChR2 in the long-range projections of paraventricular nucleus of the hypothalamus (PVN) neurons to brain stem cardiac vagal neurons (CVNs). Using PVN subpopulation-specific promoters for vasopressin and oxytocin, we were able to depolarize the soma of these neurons upon photostimulation, but these promoters were not strong enough to drive sufficient expression for optogenetic stimulation and synaptic release from the distal axons. However, utilizing the synapsin promoter photostimulation of distal PVN axons successfully evoked glutamatergic excitatory post-synaptic currents in CVNs. Employing the Cre/loxP system, using the Sim-1 Cre-driver mouse line, we found that the Rosa-CAG-LSL-ChR2-EYFP Cre-responder mice expressed higher levels of ChR2 than the Rosa-CAG-LSL-ChR2-tdTomato line in the PVN, judged by photo-evoked currents at the soma. However, neither was able to drive sufficient expression to observe and photostimulate the long-range projections to brainstem autonomic regions. We conclude that a viral vector approach with a strong promoter is required for successful optogenetic stimulation of distal axons to evoke transmitter release in pre-autonomic PVN neurons. This approach can be very useful to study important hypothalamus-brainstem connections, and can be easily modified to selectively activate other long-range projections within the brain.

Oxytocin promotes functional coupling between paraventricular nucleus and both sympathetic and parasympathetic cardioregulatory nuclei

“…Based on our correlational approach, functional coupling between the PVN and brainstem cardioregulatory regions was apparently not driven by activated cells synthesizing OXT, thus suggesting the involvement of other systems. For example, these findings may be consistent with recent work demonstrating the involvement of glutamate receptors in mediating downstream effects of OXT-receptor activation (Piñol et al, 2012). …”

“…AVP, corticotropin releasing hormone) that is critical to the stress buffering and anxiolytic properties of OXT (Smith et al, 2015; Smith and Wang, 2014). The PVN functions in coordination with specific brainstem nuclei known to regulate the autonomic nervous system, including the dorsal vagal complex (DVC), nucleus ambiguous (AMB), and rostral ventrolateral medulla (RVLM) (Herman et al, 1996; Piñol et al, 2012). Plasma corticosterone concentrations provided an index of the functional output of the HPA axis.…”

Neurotransmission to parasympathetic cardiac vagal neurons in the brain stem is altered with left ventricular hypertrophy-induced heart failure

“…These four populations of GABAergic neurons were retained in the brain stem slice preparation used in this study and are the probable source of the GABAergic neurons that directly project to CVNs that are facilitated in LVH animals. There are two known major sources of excitatory input to CVNs: one originates from neurons in the NTS (27), and the other is an excitatory pathway from neurons in the hypothalamic paraventricular nucleus (PVN) (9,29,30). The NTS receives primary information from cardiorespiratory sensory neurons, and the excitatory pathway from the NTS to CVNs likely plays an essential role in the chemoreceptor and baroreceptor reflex control of heart rate (1).…”

Translational and therapeutic potential of oxytocin as an anti-obesity strategy: Insights from rodents, nonhuman primates and humans

“…Optogenetic stimulation of PVN OT neurons suppresses the feeding response evoked by photostimulation of orexigenic agouti-related protein (AGRP) neurons, but photostimulation of PVN OT neurons alone fails to inhibit refeeding following a 24-h fast in mice [114]. While it will be important to determine if optogenetic activation of PVN OT neurons is sufficient to elicit release of OT from long axonal projections to the hindbrain [115,116], these findings are, however, consistent with what appears to be a relatively minor role of PVN OT neurons to suppress the refeeding response following a prolonged fast in rats [117]. Shah and colleagues demonstrated that reexpression of MC4Rs in PVN OT neurons in oxytocin-Ires-Cre;Mc4R loxTB / loxTB mice failed to attenuate the obesity phenotype in MC4R null MC4R loxTB / loxTB mice [118].…”