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. Author manuscript; available in PMC: 2022 Jun 14.
Published in final edited form as: Sci Transl Med. 2021 Nov 24;13(621):eabl4580. doi: 10.1126/scitranslmed.abl4580

Fig. 3. Hcn2R/R mice have less Ih and more temporal summation in the TA pathway.

Fig. 3.

(A) Representative Western blots of HCN1, HCN2, or TRIP8b, quantified in (B) (HCN1 Hcn2+/+: 100.0 ± 6.7; Hcn2R/R: 94.4 ± 8.2, n = 3 and 6; HCN2 Hcn2+/+: 100.0 ± 4.4; Hcn2R/R: 98.5 ± 6.4, n = 3 and 5; TRIP8b Hcn2+/+: 100.0 ± 8.1; Hcn2R/R: 97.8 ± 3.5, n = 3 and 5). Uncropped blots are provided in fig. S7. (C) Representative images of CA1 stained with anti-HCN2 antibody. Scale bar, 100 μm. (D) Quantification of HCN2 staining [Hcn2+/+: 881.8 ± 24.1 arbitrary units (AU); Hcn2R/R: 850.5 ± 41.1 AU, n = 4 and 4]. (E) Representative current clamp recordings showing response to −100-pA current injection from a −70-mV potential. Note that the y axis is scaled to maximal deflection to facilitate comparison (see Materials and Methods). (F) Quantification of sag ratio (Hcn2+/+: 1.38 ± 0.02; Hcn2R/R: 1.28 ± 0.03, n = 8 and 8, N = 5 and 4). (G) Half activation potential for Ih in the presence of 8-Br-cAMP (Hcn2+/+: −84.3 ± 1.1 mV; Hcn2R/R: −94.2 ± 1.7 mV, n = 8 and 8, N = 5 and 4). (H) Membrane resistance at −60 mV in the presence or absence of 8-Br-cAMP (Hcn2+/+, pre–8-Br-cAMP: 242.5 ± 30.2 megohms; Hcn2+/+, post–8-Br-cAMP: 242.0 ± 19.6 megohms; Hcn2R/R, pre–8-Br-cAMP: 257.6 ± 17.2 megohms; Hcn2R/R, post–8-Br-cAMP: 347.2 ± 20.4 megohms; n = 8 and 8, N = 5 and 4). (I) Action potentials produced in response to current injection in the absence of 8-Br-cAMP (left) or the presence of 8-Br-cAMP (right). (J) Recording configuration for examining the TA pathway. Whole-cell recording was performed from CA1 cell body (green), whereas bipolar simulating electrode was placed on the TA pathway (in blue). (K) Representative traces obtained from the configuration in (J) showing temporal summation before and after application of ZD7288. (L) Quantification of the change in temporal summation (change in EPSP5/EPSP1 with ZD7288 Hcn2+/+: 0.97 ± 0.34 mV; Hcn2R/R: 0.11 ± 0.16 mV, n = 4 and 7, N = 4 and 5). *P < 0.05. SO, stratum oriens; SP, stratum pyramidale; SR, stratum radiatum; SLM, stratum lacunosum moleculare.