ACE2 expression on inhibitory neurons controls neuronal excitability of the PVN
Authors: Mukerjee Snigdha , Gao Hong , Zsombok Andrea , Lazartigues Eric 
Published: June 2021

ACE2 is a compensatory enzyme converting Angiotensin (Ang)‐II to Ang‐(1–7). ACE2 is expressed on neurons in the brain, including in the paraventricular nucleus (PVN) that contains pre‐sympathetic neurons and is a major source of elevated sympathetic outflow. Lack of ACE2 on the cell membrane contributes to neurogenic hypertension in experimental models and patients while ACE2 over expression normalizes autonomic and baroreflex functions. We hypothesize that ACE2 maintains inhibitory control of the PVN to impede sympathetic overdrive and prevent excessive neuronal excitability.

To characterize the type of neurons that express ACE2, hypothalamic primary neurons cultures from C57Bl6/J mouse neonates (1–2 days old) were maintained for 10 days before immunolabeling. ACE2 was shown to co‐localize with inhibitory GABAergic neurons (GAD67) and oxytocinergic neurons, with intense cell body labeling, but also with vasopressinergic neurons. Interestingly, ACE2 did not co‐localize with excitatory glutamatergic neurons but these vGluT2‐labeled cells were shown to synapse onto ACE2 expressing neurons. To verify the lack of ACE2 expression on glutamatergic PVN neurons, we used transgenic mice where vGluT2‐driven cre recombinase allows expression of a tdTomato reporter. Immuno‐labeling of PVN brain sections with ACE2 confirmed the absence of the enzyme on these excitatory neurons.

To clarify the role of ACE2 on PVN neuronal excitability, we generated a transgenic mouse (ACN) where the floxed ACE2 gene is deleted from neurons, using CreLoxP technology (synapsin‐cre). To identify blood pressure (BP)‐relevant neurons in the PVN, a GFP‐tagged pseudorabies virus (PRV‐152) was injected in the kidney of male ACN and control (CT) mice. After 4.5 days, whole‐cell patch clamp recording was performed on PVN kidney‐related neurons. GFP‐tagged cells in the PVN showed a low‐threshold spike, characteristic of pre‐autonomic neurons. ACE2 deletion did not change resting membrane potential or excitatory post‐synaptic currents. Although, the frequency of spontaneous inhibitory post‐synaptic currents (IPSC) was unchanged, their amplitude was significantly reduced (ACN: 13.7 ±1 pA, n=14 vs. CT: 19.2 ±3 pA, n=8; Student's t test p<0.05). Moreover, tetrodotoxin application revealed a change in unprompted events in the absence of presynaptic action potentials. Lack of ACE2 was associated with reduced frequency (ACN: 0.8 ±0.2 Hz, n=15 vs. CT: 1.5 ±0.3 Hz, n=9; t test p<0.05) and amplitude (ACN: 12.5 ±1 pA, n=15 vs. CT: 16.1 ±2 pA, n=9; t test p<0.0001) of miniature IPSC, indicating decreased inhibitory neurotransmitter release or altered post‐synaptic mechanisms in ACN mice.

Together, our data show that ACE2 is prevalent on inhibitory neurons synapsing to PVN neurons involved in BP regulation. The presence of ACE2 might sensitize the PVN to inhibitory inputs, therefore restraining glutamatergic neurons from over‐excitation and a potential sympathetic overdrive. As loss of ACE2 is associated with increased systolic pressure in patients, manipulation of “ACE2 neuron” activity provides a new therapeutic approach for the treatment of hypertension. Next, we will clarify the role of ACE2 on vasopressinergic neurons.

Support or Funding Information

NIH/NHLBI HL093178

COBRE P30GM106392

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.