Sympathoexcitation by hypothalamic paraventricular nucleus neurons projecting to the rostral ventrolateral medulla

By Satoshi Koba, Eri Hanai, Nao Kumada,  Naoya Kataoka, Kazuhiro Nakamura, and Tatsuo Watanabe 

Division of Integrative Physiology, Tottori University Faculty of Medicine, 86 Nishi-cho, Yonago, Tottori 683-8503, JapanDepartment of Integrative Physiology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan PRESTO, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012, Japan 
Journal of Physiology, 596.19 (2018) pp 4581–4595 

The rostral ventrolateral medulla (RVLM), a brain region involved in the baroreceptor reflex, is essential in the regulation of blood pressure through spinally projecting neurons that contribute to the sympathetic nervous output. The RVLM receives afferent input from the paraventricular nucleus (PVN), which is thought to regulate the outflow from the RVLM (PVN-RVLM neurons). However, the excitatory role of the PVN-RVLM neurons has never been researched prior to this study. Through the use of optogenetics, Koba et al. investigated the excitatory role of the PVN-RVLM neurons on the effects of renal sympathetic nerve activity (RSNA) and the coinciding changes in mean arterial pressure (MAP).

Three experiments were completed in order to answer the study’s question: 1. Photostimulation of PVN-RVLM axons on renal sympathetic nerve activity; 2. The effects of glutamate receptor blockade in the RVLM on the photostimulation of PVN and; 3. The effects of intermittent photostimulation of PVN-RVLM neuronal cell bodies on renal sympathetic nerve activity.  

The study used rats more than 7 weeks old to complete the experiments. To prepare the animals for the optogenetic studies, their PVN-RVLM neurons were first transfected with either a control virus vector (pAAV2-CMV-palGFP) or the channelrhodopsin-variant containing vector (pAAV2-CMV-ChIEF-tdTomato). The vectors were microinjected into either the PVN unilaterally or the RVLM bilaterally, depending on which experiment the animals were used for. Immunofluorescence staining determined which type of neuron was transfected after the experiment.

Experiment 1 animals received unilateral microinjections of the vectors unilaterally within the PVN. The experiment found that the active vector animals had significant increases in RSNA and MAP compared to baseline after photostimulation within the RVLM. Thus, the pre-synaptic PVN neurons expressed channelrhodopsin, which would activate the neurons at PNV-RVLM neurons, leading to excitation down to the renal sympathetic nerve. This did not happen in the control animals, signifying that they did not have channelrhodopsin expressed in the synapses of the PVN-RVLM animals. These neuron groups also expressed a large amount of VGLUT2 (vesicular glutamate transporter 2) in the PVN-derived axons, which lead the researchers to think that the glutamate transmission to the RVLM leads to excitation.

Experiment 2 investigated whether or not the PVN-RVLM neurons acted through glutamate to produce sympathoexcitation. Glutamate blockers AP5 and CNQX were unilaterally microinjected into the RVLM. After glutamate injection, the photostimulation did not produce the sympathoexcitation that was previously measured. Thus, the researchers concluded that glutamate was at least one of the neurotransmitters needed for sympathoexcitation with the PVN-RVLM neurons.

To further understand the how the PVN-RVLM neurons work, the cell bodies within the PVN received photostimulation, rather than the axons (Exp. 1). The cell bodies received 1-minute intervals of photostimulation at 10, 20, and 40 Hz, revealing a “synchronous” activation at the level of the renal sympathetic nerve. The level of activation correlated to the level of Hz used to stimulate the nerve.

The researchers concluded that the glutaminergic PVN-RVLM neurons help to drive sympathoexcitation. These neurons act on C1 neurons within the RVLM, which then project down the spinal cord to modulate blood pressure in the rats. While this study was interesting, it stuggled with transfecting all of the animals. Therefore,  it is difficult to say that all possible PVN-RVLM neurons were correctly transfected, which may produce inaccurate results otherwise. More research should be done of the efficacy of the optogenetic methods used within this paper. Furthermore, the role of non-C1 neurons, which also receive PVN input, may have been activated as well. Their activation may play a role in sympathoexcitation, and more research should be done. It would also be interesting to see the effects in rats younger than 7 weeks old. Additionally, female rats should be investigated, due to effects that estrogen may have on baroreceptor reflex. 

-LivInLaVida