Chronic Treatment of a Mouse Model of Cerebral Amyloid Angiopathy and Brain AT1 Receptor Expression

Abstract

Introduction: The renin-angiotensin-aldosterone system (RAAS) has been shown to be dysregulated in dementia, with elevated levels of angiotensin-converting enzyme (ACE), angiotensin (Ang) II, and Ang II type 1 receptors (AT₁Rs). Cerebral amyloid angiopathy (CAA), a common cerebrovascular disease, currently has no treatment or cure available. We aimed to determine if a mouse model with CAA (Tg-SwDI) also exhibits elevated levels of AT₁Rs and whether RAAS-targeting drugs (telmisartan and lisinopril) mitigate these effects.

Materials and methods: Tg-SwDI mice were treated with sub-depressor doses of either telmisartan or lisinopril from 3-8 months of age, with blood pressure being monitored 2 and 4 months after the start of treatment. Postmortem, receptor autoradiography was performed to determine levels of AT₁R in 13 brain regions in untreated and treated Tg-SwDI mice compared to wild-type controls (C57Bl/6J).

Results: No statistically significant differences among groups were observed in any of the 13 regions analyzed. However, trends with medium to large effect sizes were observed.

Conclusions: CAA did not significantly dysregulate AT₁R levels in the brains of Tg-SwDI mice compared to wild-type mice. Drug treatment caused no significant brain AT₁R alterations. Further studies are required to determine if the trends observed are pathophysiological and pharmacologically significant.

Keywords: Cerebral amyloid angiopathy; Tg-SwDI mice; lisinopril; receptor autoradiography; telmisartan.

Figure 1.. Simplified representation of the RAAS activity in the vasculature and in the brain.

(A) In the vasculature, the kidneys release the enzyme renin, along with its precursor prorenin. Renin is responsible for cleaving angiotensinogen, giving rise to angiotensin (Ang) I, which is then cleaved by the angiotensin-converting enzyme (ACE), generating Ang II. In turn, Ang II can activate both Ang II type 1 receptors (AT₁Rs) and Ang II type 2 receptors (AT₂Rs). Additionally, Ang II can be further cleaved by ACE-2, prolylcarboxypeptidase (PRCP), or carboxypeptidase A (CPA) to generate Ang-(1–7), which can activate MasRs and AT₂Rs (to a lesser extent), and inhibit AT₁Rs, counteracting the effects of Ang II. Alternatively, Ang-(1–7) can also be formed through cleavage of Ang I by different enzymes. (B) In the brain, astroglia produce angiotensinogen, which can be cleaved by renin, prorenin, or a renin-like enzyme present in neurons, to give rise to Ang I^, . In turn, ACEs, which are expressed on the cell membranes of the brain, cleave Ang I, giving rise to Ang II. Like in the vasculature, Ang-(1–7) activity in the brain counteracts the effects of Ang II. Created in BioRender. Speth, R. (2025) https://BioRender.com/rzfj0v4

Figure 2.. Generic timeline of the study for each cohort.

Mice started on the drug treatment at approximately 3 months of age, undergoing blood pressure measurements 2 and 4 months into treatment. Euthanasia occurred at approximately 8 months of age, when significant CAA pathology is observed in Tg-SwDI mice, followed by cryostat sectioning of brains and RAR. Created in BioRender. Speth, R. (2025) https://BioRender.com/t7lrdza

Figure 3.. RAR experimental design.

Four Coplin jars that can hold 10 slides each were filled with AM5 and used to carry out the pre-incubation step. After 10 min in solution, slides were immediately transferred to their respective incubation jars, where they incubated for 60 min at room temperature. Each incubation jar contained 35 mL of a stock solution prepared by mixing assay medium 5 (AM5) solution (150 mM sodium chloride, 5 mM disodium EDTA, 0.1 mM Bacitracin, and 500 mM dibasic sodium phosphate; pH 7.1 – 7.2) and ¹²⁵I-SI Ang II ligand to achieve a concentration of 255 pM. Additionally, 35 μL of 10 μM losartan were added to half of the Coplin jars. Slides were blotted using a paper towel before the first two water rinses and after the last two water rinses. Each water rinse swirl did not last for more than 1 or 2 seconds. Created in BioRender. Speth, R. (2025) https://BioRender.com/omwatj2

Figure 4.. Drug intake and systolic BP.

(A) There was no statistically significant difference observed between actual drug intake and the target doses of each drug (red dotted line). In the males, p=0.3333 for telmisartan and p=0.1978 for lisinopril. In the females, p=0.4228 for telmisartan and p=0.7805 for lisinopril. (B) Systolic blood pressure was measured using the tail cuff method at 2-months post-initiation of treatment. There was a main effect of group [F (3, 99) = 16.50, p<0.0001, η_p²=0.3333]; overall, untreated Tg-SwDI (p<0.0001, d=1.187), telmisartan-treated Tg-SwDI (p<0.0001, d=1.474), and lisinopril-treated Tg-SwDI (p<0.0001, d=1.521) all had significantly lower systolic BP compared to WT mice groups. In the males, systolic BP of WTs was significantly higher than that of telmisartan-treated Tg-SwDI mice (p=0.0214, d=1.076). In the females, systolic BP of WTs was significantly higher than that of untreated Tg-SwDI mice (p<0.0001, d=2.148), telmisartan-treated Tg-SwDI mice (p<0.0001, d=1.972), and lisinopril-treated Tg-SwDI mice (p<0.0001, d=3.106). A statistically significant main effect of sex was observed [F (1, 99) = 36.50, p<0.0001, η_p²=0.2694], with multiple comparisons showing significant sex differences within the untreated Tg-SwDI mice (p=0.0001, d=1.512), telmisartan-treated Tg-SwDI mice (p=0.0079, d=0.8411), and lisinopril-treated Tg-SwDI mice (p<0.0001, d=2.413). (C) At 4 months post-initiation of treatment, statistically significant main effect of group was observed [F (3, 98) = 4.568, p=0.0049, η_p²=0.1227]; overall, lisinopril-treated Tg-SwDI mice had significantly lower systolic BP compared to WT mice groups (p=0.0025, d=0.9354). No statistically significant differences were observed between the male groups. In the females, lisinopril-treated Tg-SwDI mice had significantly lower systolic BP than WTs (p=0.0245, d=1.269). A statistically significant main effect of sex was observed [F (1, 98) = 35.62, p<0.0001, η_p²=0.2665], with significant multiple comparison differences observed within the WTs (p=0.0044, d=1.246), untreated Tg-SwDI mice (p=0.0063, d=0.9053), telmisartan-treated Tg-SwDI mice (p=0.0044, d=1.135), and lisinopril-treated Tg-SwDI mice (p=0.0013, d=1.522).

Figure 5.. Specific binding of ¹²⁵I-SI Ang II in the peri-hypothalamic regions of the brain of Tg-SwDI mice.

(A) A medium effect of treatment was observed in the PeVN, although not statistically significant [F (3, 30) = 1.538, p=0.2251, η_p²=0.1333] (N=38; no outliers). Untreated Tg-SwDI mice showed elevated concentration of AT₁R compared to WTs in both males (p=0.3966, d=0.8880) and females (p=0.9524, d=0.5083). In the males, telmisartan reduced specific binding in Tg-SwDI mice (p=0.3754, d=0.8593). No main effect of sex was observed [F (1, 30) = 0.799, p=0.379, η_p²=0.026]. (B) No main effect of treatment was observed in the PVN [F (3, 30) = 0.4686, p=0.7064, η_p²=0.0448] (N=38; no outliers). Untreated Tg-SwDI males showed elevated concentration of AT₁R compared to WTs (p=0.8405, d=0.5506). In the females, telmisartan reduced specific binding in drug-treated Tg-SwDI mice (p=0.8463, d=0.5086). No main effect of sex was observed [F (1, 30) = 1.40E-04, p=0.991, η_p²=4.67E-06]. (C) A medium effect of treatment was observed in the SFO, although not statistically significant [F (3, 24) = 1.123, p=0.3595, η_p²=0.1231] (N=32; 4 outliers removed). Untreated Tg-SwDI females showed elevated concentration of AT₁R compared to WTs (p=0.875, d=0.533). In the males, lisinopril reduced specific binding in drug-treated Tg-SwDI mice (p=0.6505, d=0.7766). In the females, lisinopril increased specific binding in drug-treated Tg-SwDI mice (p=0.9389, d=0.5023). No main effect of sex was observed [F (1, 24) = 0.4309, p=0.5178, η_p²=0.0176]. (D) No main effect of treatment was observed in the MnPO [F (3, 19) = 1.070, p=0.3852, η_p²=0.1446] (N=27; no outliers). Untreated Tg-SwDI females showed lower concentration of AT₁R compared to WTs (p=0.939, d=0.504). Telmisartan increased specific binding, compared to untreated Tg-SwDI mice, in males (p=0.3646, d=1.122) and females (p=0.7840, d=0.7719). No main effect of sex was observed [F (1, 19) = 0.1261, p=0.7264, η_p²=0.0066]. (E) No main effect of treatment was observed in the DMH [F (3, 28) = 0.3915, p=0.7601, η_p²=0.0403] (N=36; 1 outlier removed). Untreated Tg-SwDI females showed elevated concentration of AT₁R compared to WTs (p=0.5016, d=0.8193). Both drugs reduced specific binding, compared to untreated Tg-SwDI females (p=0.7085, d=1.042 for telmisartan and p=0.7242, d=0.7965 for lisinopril). A medium effect of sex was observed, although not statistically significant [F (1, 28) = 2.937, p=0.0976, η_p² =0.0949]. (F) A medium effect of treatment was observed in the OVLT, although not statistically significant [F (3, 20) = 0.6746, p=0.5777, η_p²=0.0919] (N=28; no outliers). Untreated Tg-SwDI males showed elevated concentration of AT₁Rs compared to WTs (p=0.8116, d=0.5932). Lisinopril reduced specific binding compared to untreated Tg-SwDI males (p=0.1320, d=1.791). Untreated Tg-SwDI females showed reduced concentration of AT₁Rs compared to WTs (p=0.6306, d=1.624). Both drugs increased specific binding, compared to untreated Tg-SwDI females (p=0.7464, d=1.296 for telmisartan and p=0.5198, d=1.090 for lisinopril). No main effect of sex was observed [F (1, 20) = 0.1747, p=0.6804, η_p²=0.0087].

Figure 6.. Specific binding of ¹²⁵I-SI Ang II in the striatum of Tg-SwDI mice.

(A) A medium effect of treatment was observed in the NAcc, although not statistically significant [F (3, 30) = 0.8254, p=0.4903, η_p²=0.0762] (N=38; no outliers). In the males, telmisartan increased specific binding compared to untreated Tg-SwDI mice (p=0.328, d=1.16). No main effect of sex was observed [F (1, 30) = 0.0040, p=0.9497, η_p²=0.0001). (B) No main effect of treatment was observed in the CPu [F (3, 30) = 0.6130, p=0.6119, η_p²=0.0578] (N=38; no outliers). In the males, telmisartan increased specific binding, compared to untreated Tg-SwDI males (p=0.5748, d=0.7090). No main effect of sex was observed [F (1, 30) = 0.149, p=0.702, η_p²=0.005].

Figure 7.. Specific binding of ¹²⁵I-SI Ang II in the amygdala of Tg-SwDI mice.

(A) No main effect of treatment was observed in the MeA [F (3, 29) = 0.4964, p=0.6877, η_p²=0.0488] (N=37; 1 outlier removed). Untreated Tg-SwDI females expressed elevated concentration of AT₁Rs, compared to WTs (p=0.4747, d=1.018). No main effect of sex was observed [F (1, 29) = 0.9795, p=0.3305, η_p²=0.0327]. (B) No main effect of treatment was observed in the CeA/BLA [F (3, 30) = 0.2351, p=0.8712, η_p²=0.0230] (N=38; no outliers). Untreated Tg-SwDI females expressed elevated concentration of AT₁Rs, compared to WTs (p=0.6727, d=0.9489). No main effect of sex was observed [F (1, 30) = 0.4594, p=0.5031, η_p²=0.0151].

Figure 8.. Specific binding of ¹²⁵I-SI Ang II in the lateral septum of Tg-SwDI mice.

A medium effect of treatment was observed in the lateral septum, although not statistically significant [F (3, 29) = 0.7532, p=0.5295, η_p²=0.0723] (N=37; 1 outlier removed). In the males, telmisartan increased specific binding, compared to untreated Tg-SwDI males (p=0.2965, d=1.777). Untreated Tg-SwDI females showed lower concentration of AT₁Rs than WTs (p=0.7225, d=0.6548). No main effect of sex was observed [F (1, 29) = 0.0834, p=0.7748, η_p² =0.0029].

Figure 9.. Specific binding of ¹²⁵I-SI Ang II in the hippocampus of Tg-SwDI mice.

A medium effect of treatment was observed in the dHpc, although not statistically significant [F (3, 28) = 1.208, p=0.3252, η_p²=0.1146] (N=36; 2 outliers removed). In the males, telmisartan decreased specific binding compared to untreated Tg-SwDI males (p=0.6194, d=1.081). Untreated Tg-SwDI females expressed slightly elevated concentration of AT₁Rs compared to WTs (p=0.8351, d=0.9635). Both drugs decreased specific binding in females, compared to untreated Tg-SwDI females (p=0.5473, d=0.9962 for telmisartan and p=0.5555, d=0.7856 for lisinopril). A medium effect of sex was observed, although not statistically significant [F (1, 28) = 2.881, p=0.1007, η_p² =0.0933].

Figure 10.. Specific binding of ¹²⁵I-SI Ang II in the sensorimotor cortex of Tg-SwDI mice.

A large effect of treatment was observed in the SMC, although not statistically significant [F (3, 23) = 1.768, p=0.1814, η_p²=0.1873] (N=31; 6 outliers removed). In the males, both drugs decreased specific binding, compared to untreated Tg-SwDI males (p=0.5952, d=1.151 for telmisartan and p=0.4333, d=1.766 for lisinopril). Untreated Tg-SwDI females expressed slightly elevated concentration of AT₁Rs, compared to WTs (p=0.7629, d=0.9847). Telmisartan decreased specific binding in the females compared to untreated Tg-SwDI females (p=0.3974, d=1.421). No main effect of sex was observed [F (1, 23) = 0.3049, p=0.5862, η_p²=0.0131].