Chronic lithium treatment induces novel patterns of pendrin localization and expression

Abstract

Prolonged lithium treatment is associated with various renal side effects and is known to induce inner medullary collecting duct (IMCD) remodeling. In animals treated with lithium, the fraction of intercalated cells (ICs), which are responsible for acid-base homeostasis, increases compared with renal principal cells (PCs). To investigate the intricacies of lithium-induced IMCD remodeling, male Sprague-Dawley rats were fed a lithium-enriched diet for 0,1, 2, 3, 6, 9, or 12 wk. Urine osmolality was decreased at 1 wk, and from 2 to 12 wk, animals were severely polyuric. After 6 wk of lithium treatment, approximately one-quarter of the cells in the initial IMCD expressed vacuolar H⁺-ATPase, an IC marker. These cells were localized in portions of the inner medulla, where ICs are not normally found. Pendrin, a Cl^-/[Formula: see text] exchanger, is normally expressed only in two IC subtypes found in the convoluted tubule, the cortical collecting duct, and the connecting tubule. At 6 wk of lithium treatment, we observed various patterns of pendrin localization and expression in the rat IMCD, including a novel phenotype wherein pendrin was coexpressed with aquaporin-4. These observations collectively suggest that renal IMCD cell plasticity may play an important role in lithium-induced IMCD remodeling.

Keywords: acid-base homeostasis; acidosis; diabetes insipidus; lithium; pendrin.

Fig. 1.

Physiological analyses of lithium-induced polyuria over 12 wk of lithium treatment. Male rats were fed a 40 mmol/kg lithium diet for 0, 1, 2, 3, 6, 9, or 12 wk. Single animals were subsequently placed in individual metabolic cages to collect urine and serum at the determined time point. A: serum lithium levels were measured using an EasyLyte according to the manufacturer’s instructions. B: urine was collected over 24 h under oil to prevent evaporation. C: urine osmolality was measured from collected urine. Data are presented as means ± SE, where *P < 0.05 vs. control. Control, n = 11; 1 wk, n = 7; 2 wk, n = 6; 3 wk, n = 8; 6 wk, n = 10; 9 wk, n = 3; 12 wk, n = 3.

Fig. 2.

Inner medullary collecting duct (IMCD) principal cells label positively for proliferating cell nuclear antigen (PCNA) during early lithium treatment. Shown are representative images from kidney inner medullary tissue (IM-3) stained for the proliferation marker PCNA (green) and the renal principal cell marker, aquaporin-4 (AQP4) (red) in untreated rats (A), 1-wk lithium-treated rats (B), 2-wk lithium-treated rats (C), and 12-wk lithium-treated rats (D). Nuclei are stained blue with DAPI; n = 3.

Fig. 3.

Inner medullary collecting duct (IMCD) principal cells are not apoptotic during lithium treatment. Shown are representative images from kidney IM-3 tissue stained for the renal principal cell marker aquaporin-4 (AQP4) (red) following a TUNEL assay (green) in untreated rats (A), 1-wk lithium-treated rats (B), 6-wk lithium-treated rats (C), and 12-wk lithium-treated rats (D). Nuclei are stained blue with DAPI; n = 3.

Fig. 4.

Chronic lithium treatment induces inner medullary collecting duct (IMCD) remodeling. Shown are representative images from kidney inner medullary (IM) tissue stained for the renal principal cell marker aquaporin-4 (AQP4) (green) and an intercalated cell marker vacuolar H⁺-ATPase (V-ATPase; red), in untreated rats (A), 1-wk lithium-treated rats (B), 2-wk lithium-treated rats (C), 3-wk lithium-treated rats (D), 6-wk lithium-treated rats (E), 9-wk lithium-treated rats (F), and 12-wk lithium-treated rats (G). H: quantification of the proportions of AQP4-positive and V-ATPase-positive cells. Data are presented as mean proportion ± SE of the proportion, where *P < 0.05 vs. control. For figure clarity, cells both AQP4 and V-ATPase positive were excluded from this analysis, but their proportion can be found in Table 2. Nuclei are stained blue with DAPI; n = 3.

Fig. 5.

Chronic lithium treatment induces aberrant patterns of intercalated cell type expression in the kidney. Shown are representative images from kidney inner medullary (IM) tissue stained for an intercalated cell marker and vacuolar H⁺-ATPase (V-ATPase) (red) and pendrin (green) in 6-wk lithium-treated rats (A), 9-wk lithium-treated rats (B), and 12-wk lithium-treated rats (C). D and E: representative images at a lower magnification of staining in untreated and 12-wk lithium-treated rats. Nuclei are stained blue with DAPI; n = 3.

Fig. 6.

Altered intercalated cell types are observed after chronic lithium treatment. Shown are representative images from kidney inner medullary (IM) tissue from 12-wk lithium-treated rats stained for an intercalated cell marker vacuolar H⁺-ATPase (V-ATPase) (red) and pendrin (green). Observed were the following cell types: non-A, non-B type intercalated cells (A); B-type intercalated cells (B), and a novel renal cell phenotype (C), wherein both pendrin and aquaporin-4 (AQP4) (red), a principal cell marker, are expressed. D: quantification proportions of different intercalated cell types. Control, n = 3; 6 wk, n = 2, 9 wk, n = 3, 12 wk, n = 2.

Fig. 7.

Acid-base homeostasis was disrupted during lithium treatment. A: urine pH was measured in urine collected under oil from rats in fed a 40 mmol/kg lithium diet for 0, 1, 2, 3, 6, 9, or 12 wk. B: urine ammonium was measured according to manufacturer’s instructions in urine collected under oil from rats fed a 40 mmol/kg lithium diet for 0, 1, 2, 3, 6, 9, or 12 wk. Data are presented as means ± SE, where *P < 0.05 vs. control. Control, n = 11; 1 wk, n = 7; 2 wk, n = 6; 3 wk, n = 8; 6 wk, n = 10; 9 wk, n = 3; 12 wk, n = 3.