Concurrent administration of thyroxine and donepezil induces plastic changes in the prefrontal cortex of adult hypothyroid rats

Abstract

The aim of the present study was to observe the effects of the concurrent administration of thyroxine (T4) and an acetylcholinesterase (AChE) inhibitor, donepezil (DON), on the hypothyroidism‑induced ultrastructural changes of the prefrontal cortex (PFC) in adult rats. The acetylcholine (ACh) content and AChE activity was assessed, as well as the expressions of synaptotagmin‑1 (syt‑1) and SNAP‑25 were analyzed in the rats. Adding 0.05% propylthiouracil to rats' drinking water induced a hypothyroid rat model. The animals were treated with T4 and DON administered separately or in combination from the fifth week. Transmission electron microscope analysis revealed that hypothyroidism induced marked ultrastructural changes, including the neurons, the synapses and the myelin sheath in the PFC. T4 or DON treatment improved the morphologic features of the PFC, and the performance of the T4 combined DON group was the closest to the control group. Moreover, hypothyroidism significantly decreased the content of ACh (29.8%) and activity of AChE (27.8%), which were restored to control values by T4 admi-nistration. In addition, DON treatment restored ACh content to normal. At the protein level, hypothyroidism increased the levels of syt‑1 and SNAP‑25 in the PFC, both of which were not restored to control values following T4 administration, while concurrent administration of T4 and DON was able to induce this effect. These results suggested that adult‑onset hypothyroidism induce morphological, biochemical and molecular alterations in the PFC, combined administration of T4 and DON induce plastic changes in the PFC, different from that of the standard T4 therapy, and that the DON treatment may facilitate the recovery of synaptic protein impairments induced by hypothyroidism.

Figure 1.

Serum T3 and T4 levels among the groups. The rats had significantly lower serum (A) T3 and (B) T4 levels in the Hypo and the DON group, compared to the rats in the CON group. The levels of T3 and T4 were restored to control values by T4 and T4 + DON treatment. The analysis was performed by analysis of variance. Data are presented as the mean ± standard error of the mean. *P<0.05 vs. CON group. T3, triiodothyronine; T4, thyroxine; Hypo, hypothyroid; DON, donepezil; CON, control; SEM, standard error of the mean.

Figure 2.

Representative images of the rat PFC neurons among the groups by TEM (magnification, ×15,000). (A) CON group; (B) Hypo group; (C) DON group; (D) T4 group; (E) T4 + DON group. PFC, prefrontal cortex; TEM, transmission electron microscope; CON, control; Hypo, hypothyroid; DON, donepezil; T4, thyroxine.

Figure 3.

Representative images of the rat PFC synaptic ultrastructures among the groups by TEM (magnification, ×40,000). (A) CON group; (B) Hypo group; (C) DON group; (D) T4 group; (E) T4 + DON group. PFC, prefrontal cortex; TEM, transmission electron microscope; CON, control; Hypo, hypothyroid; DON, donepezil; T4, thyroxine.

Figure 4.

Representative images of the rat PFC myelin sheath among the groups by TEM (magnification, ×20,000). (A) CON group; (B) Hypo group; (C) DON group; (D) T4 group; (E) T4 + DON group. PFC, prefrontal cortex; TEM, transmission electron microscope; CON, control; Hypo, hypothyroid; DON, donepezil; T4, thyroxine.

Figure 5.

Concentration of prefrontal ACh from CON, Hypo, DON, T4 and T4 + DON groups (n=10), homogenates were extracted from the PFC of each rat. Data are presented as means ± standard error of the mean of three independent experiments. *P<0.05 vs. CON. ACh, acetylcholine; T4, thyroxine; DON, donepezil; CON, control; Hypo, hypothyroid.

Figure 6.

The AChE activity in the PFC of rats from CON, Hypo, DON, T4 and T4 + DON groups (n=10), homogenates were extracted from the PFC of each rat. Data are presented as means ± standard error of the mean of three independent experiments. *P<0.05 vs. CON. CON, control; Hypo, hypothyroid; T4, thyroxine; DON, donepezil; AChE, acetylcholinesterase; Prot, prefrontal protein.

Figure 7.

T4 and DON regulated the expression of syt-1 and SNAP-25 protein in the PFC of adult hypothyroid rats. (A and B) Western blot analysis of the expressions of syt-1 and SNAP-25 protein in the PFC of CON, Hypo, DON, T4 and T4 + DON groups (C and D). Quantification analysis of the relative protein levels in each group. The data are presented as the means ± standard error of the mean (n=10). *P<0.05 vs. CON. CON, control; Hypo, hypothyroid; DON, donepezil; T4, thyroxine; syt-1, synaptotagmin-1; SNAP-25, synaptosomal-associated protein of 25 kDa; PFC, prefrontal cortex; OD, optical density.