Tyrosine Hydroxylase‐Positive Neurons in the Hypothalamic Paraventricular Nucleus Modulate Blood Glucose Levels in Mice

Abstract

Tyrosine hydroxylase (TH)‐positive neurons in the paraventricular nucleus of the hypothalamus (PVN), termed THPVN neurons, have emerged as key metabolic regulators. Recent studies showed that THPVN neurons are mostly Gamma‐aminobutyric acid (GABA)ergic and/or dopaminergic. Previous results from our laboratory indicate that THPVN neurons project to brainstem autonomic regulatory nuclei, such as nucleus tractus solitarius (NTS) and rostral ventrolateral medulla (RVLM). However, it is not known if THPVN neurons play a role in blood glucose regulation. We hypothesize that THPVN neurons are key regulators of blood glucose. To test our hypothesis, we used adeno‐associated virus serotype 2 (AAV2)‐mediated Cre‐recombinase driven by a rat TH promoter (AAV2‐TH‐Cre), in combination with chemogenetic approach (Designer Receptors Exclusively Activated by Designer Drugs, DREADD) to activate or inhibit THPVN neurons and continuously monitored blood glucose levels using telemetry system. hM3Dq‐LoxP (excitatory‐DREADD) and hM4Di‐LoxP (inhibitory‐DREADD) mice, 10‐14 weeks old, were bilaterally microinjected with AAV2‐TH‐Cre into the PVN. One month following AAV injection, mice were implanted with telemetry glucose transmitters (HD‐XG, DSI) to monitor blood glucose (BG) levels. To activate or inhibit THPVN neuronal activity, Clozapine‐N‐oxide (CNO, 1 mg/Kg) or vehicle were administered (i.p.) and BG was monitored by telemetry. To examine the functional role of THPVN neurons at baseline and avoid the influence of feeding on blood glucose levels, mice were fasted for 2 hours and fasting was maintained throughout the experiment. We found that activation of THPVN neurons by CNO in hM3Dq‐LoxP mice did not alter baseline blood glucose (n=5); on the other hand, inhibition of THPVN neurons by CNO in hM4Di‐LoxP mice increased baseline blood glucose levels compared with vehicle only in male mice (Male: ΔBG: 31.6 ± 8.6 mg/dL vs. 11.0 ± 6.7 mg/dL, n=4, p=0.03; Female: ΔBG: 11.0 ± 7.4 mg/dL vs. 7.3 ± 5.9 mg/dL, n=4, p=0.16). To study the functional role of THPVN neurons on whole body glucose handling, mice were fasted overnight and simultaneously injected with glucose (1 g/Kg) and either vehicle or CNO. We found that activation of THPVN neurons by CNO improved glucose clearance after glucose overload when compared with vehicle (AUC: 3133.0±109.0 vs. 6021.0±102.0, p<0.0001, n=3) in hM3Dq‐LoxP mice. In summary, our data suggest that THPVN neurons negatively regulate blood glucose possibly by their GABAergic input to downstream neural circuits. We conclude that THPVN neurons play a functional role in the regulation of glucose homeostasis; while the mechanisms remain to be determined.