Metoprolol prevents neuronal dendrite remodeling in a canine model of chronic obstructive sleep apnea

Abstract

Obstructive sleep apnea (OSA) is closely associated with central nervous system diseases and could lead to autonomic nerve dysfunction, which is often seen in neurodegenerative diseases. Previous studies have shown that metoprolol prevents several chronic OSA-induced cardiovascular diseases through inhibiting autonomic nerve hyperactivity. It remains unclear whether chronic OSA can lead to dendritic remodeling in the brain, and whether metoprolol affects the dendritic remodeling. In this study we investigated the effect of metoprolol on dendrite morphology in a canine model of chronic OSA, which was established in beagles through clamping and reopening the endotracheal tube for 4 h every other day for 12 weeks. OSA beagles were administered metoprolol (5 mg· kg^-1· d^-1). The dendritic number, length, crossings and spine density of neurons in hippocampi and prefrontal cortices were assessed by Golgi staining. And the protein levels of hypoxia-inducible factor-1α (HIF-1α) and brain-derived neurotrophic factor (BDNF) were measured by Western blotting. We showed that chronic OSA successfully induced significant brain hypoxia evidenced by increased HIF-1α levels in CA1 region and dentate gyrus of hippocampi, as well as in prefrontal cortex. Furthermore, OSA led to markedly decreased dendrite number, length and intersections, spine loss as well as reduced BDNF levels. Administration of metoprolol effectively prevented the dendritic remodeling and spine loss induced by chronic OSA. In addition, administration of metoprolol reversed the decreased BDNF, which might be associated with the metoprolol-induced neuronal protection. In conclusion, metoprolol protects against neuronal dendritic remodeling in hippocampi and prefrontal cortices induced by chronic OSA in canine.

Keywords: BDNF; HIF-1α; canine model; chronic obstructive sleep apnea; hippocampus; metoprolol; neuronal dendritic remodeling; prefrontal cortex.

Fig. 1. Construction of the OSA canine model.

a Schematic diagram of the chronic OSA beagle model. At the first week, the apnea AHI was set as 6. The duration of trachea blockage was 1 min, and ventilation was 9 min. In the next 3 weeks, the ventilation duration was progressively 1 min shorter than the previous week. In the last 8 weeks, the duration of tracheal ventilation was 5 min, and the AHI was 10. The ventilation and blockage were exchanged for 4 h every other day in the whole process. b Negative intra-airway pressure at baseline and apnea for 30 and 60 s. **P < 0.01 vs. baseline, n = 4 for each group, and each value represents the mean ± SEM. c HIF-1α expression level in the hippocampal CA1 and DG as well as in the prefrontal cortex (for CA1: n = 7; DG: n = 5; prefrontal cortex: n = 5, with four beagles per group). **P < 0.01 vs. sham control group, values are expressed as the mean ± SEM.

Fig. 2. Metoprolol rescues the degeneration of pyramidal neurons in the hippocampal CA1 of the OSA canine model.

a Representative photomicrograph (left) and tracing image (right) of pyramidal neurons in the hippocampal CA1 from the sham control group (upper), OSA group (middle) and OSA treated with metoprolol group (below), scale bar = 100 μm. b−g Quantification of total dendrite length (b), total dendrite number (c), apical dendrite length (d), basal dendrite length (e), apical dendrite number (f), and basal dendrite number (g) in hippocampal pyramidal neurons. h Schematic experimental diagram of Sholl analysis. i−k Sholl analysis of the intersection number of total dendrites (10–200 μm) (i), apical dendrites (10–200 μm) (j) and basal dendrites (10–120 μm) (k). *P < 0.05, **P < 0.01, ***P < 0.001 vs. sham control group, ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 vs. OSA group, values are expressed as the mean ± SEM.

Fig. 3. Metoprolol reverses the degeneration of prefrontal pyramidal neurons in the OSA canine model.

a Representative photomicrograph (left) and tracing image (right) of pyramidal neurons in prefrontal cortices from the sham control group (top), OSA group (middle) and OSA with metoprolol treatment group (bottom), scale bar = 100 μm. b−g Quantification of prefrontal pyramidal overall dendrite length (b), overall dendrite number (c), apical dendrite length (d), basal dendrite length (e), apical dendrite number (f), and basal dendrite number (g). h−j Sholl analysis of the intersection number of total dendrites (10–200 μm) (h), apical dendrites (10–200 μm) (i) and basal dendrites (10–120 μm) (j). *P < 0.05, **P < 0.01, ***P < 0.001 vs. sham control group, ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 vs. OSA group, values are expressed as the mean ± SEM.

Fig. 4. Metoprolol alleviates granular cell degeneration in the hippocampal DG and nonpyramidal neuron regression in the prefrontal cortex of the OSA model.

a Representative photomicrograph (left) and tracing image (right) of granular neurons of the hippocampal DG region from the sham control group (left), OSA group (middle) and OSA treated with metoprolol group (right), scale bar = 50 μm. b, c Quantification of the dendrite length (b) and dendrite number (c) of hippocampal granular cells. d Representative photomicrograph (left) and tracing image (right) of nonpyramidal neurons in the prefrontal cortex. e, f Quantification of the dendrite length (e) and dendrite number (f) of prefrontal nonpyramidal cells. *P < 0.05, **P < 0.01, ***P < 0.001 vs. sham control group, ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 vs. OSA group, values are expressed as the mean ± SEM.

Fig. 5. Metoprolol ameliorates the reduction in dendritic spine density in the OSA canine model.

a−d Representative photomicrograph schematic diagram of selected dendritic spines of the apical dendrites (a) and basal dendrites (b) of hippocampal and prefrontal cortical apical dendrites (c) and basal dendrites (d) from the sham control group (left), OSA group (middle) and OSA treated with metoprolol group (right), scale bar = 5 μm. e, f Quantification of apical (e) and basal (f) dendritic spine density in hippocampal dendrites. g, h Quantification of apical (g) and basal (h) dendritic spine density of prefrontal cortical dendrites. *P < 0.05, ***P < 0.001 vs. sham control group, ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 vs. OSA group, values are expressed as the mean ± SEM.

Fig. 6. BDNF level in the hippocampi and prefrontal cortices of sham, OSA and metoprolol-treated OSA beagles.

a−c BDNF expression level in the hippocampal CA1 (a) (n = 4, with four beagles per group) and hippocampal DG regions (b) (n = 5, with four beagles per group) and the prefrontal cortex (c) (n = 5, with four beagles per group) from the sham group, OSA group and OSA treated with metoprolol group. **P < 0.01, ***P < 0.001 vs. sham control group, ^##P < 0.01 vs. OSA group, values are expressed as the mean ± SEM.