Trabecular bone loss after administration of the second-generation antipsychotic risperidone is independent of weight gain

Abstract

Second generation antipsychotics (SGAs) have been linked to metabolic and bone disorders in clinical studies, but the mechanisms of these side effects remain unclear. Additionally, no studies have examined whether SGAs cause bone loss in mice. Using in vivo and in vitro modeling we examined the effects of risperidone, the most commonly prescribed SGA, on bone in C57BL6/J (B6) mice. Mice were treated with risperidone orally by food supplementation at a dose of 1.25 mg/kg daily for 5 and 8 weeks, starting at 3.5 weeks of age. Risperidone reduced trabecular BV/TV, trabecular number and percent cortical area. Trabecular histomorphometry demonstrated increased resorption parameters, with no change in osteoblast number or function. Risperidone also altered adipose tissue distribution such that white adipose tissue mass was reduced and liver had significantly higher lipid infiltration. Next, in order to tightly control risperidone exposure, we administered risperidone by chronic subcutaneous infusion with osmotic minipumps (0.5 mg/kg daily for 4 weeks) in 7 week old female B6 mice. Similar trabecular and cortical bone differences were observed compared to the orally treated groups (reduced trabecular BV/TV, and connectivity density, and reduced percent cortical area) with no change in body mass, percent body fat, glucose tolerance or insulin sensitivity. Unlike in orally treated mice, risperidone infusion reduced bone formation parameters (serum P1NP, MAR and BFR/BV). Resorption parameters were elevated, but this increase did not reach statistical significance. To determine if risperidone could directly affect bone cells, primary bone marrow cells were cultured with osteoclast or osteoblast differentiation media. Risperidone was added to culture medium in clinically relevant doses of 0, 2.5 or 25 ng/ml. The number of osteoclasts was significantly increased by addition in vitro of risperidone while osteoblast differentiation was not altered. These studies indicate that risperidone treatment can have negative skeletal consequences by direct activation of osteoclast activity and by indirect non-cell autonomous mechanisms. Our findings further support the tenet that the negative side effects of SGAs on bone mass should be considered when weighing potential risks and benefits, especially in children and adolescents who have not yet reached peak bone mass.

Figure 1

Reduced body fat and femur areal bone mineral density after risperidone administration. Risperidone was administered to male B6 mice orally in the food for five or eight weeks, starting at 3.5 weeks of age. Lean mass, fat mass, total and femur areal BMD were measured by DXA. Bars represent mean ± standard error of untreated (white) and risperidone treated mice (black). N=8. *p<0.05 by Student’s t-test.

Figure 2

Reduced trabecular and cortical bone mass in the femur after eight weeks of risperidone administration. Shown are representative three-dimensional µCT images from control and risperidone treated mice.

Figure 3

Reduced white adipose tissue and increased liver adiposity after five weeks of risperidone treatment. (A) Representative abdominal MR images and epididymal fat mass. (B) Liver Oil Red O staining (red stain indicates presence of lipids) and liver mass. Bars represent mean ± standard error of untreated (white) and risperidone treated mice (black). N=8. *p<0.05 by Student’s t-test.

Figure 4

Subcutaneous infusion of risperidone did not alter total body, lean or fat mass, but did reduce femur areal BMD and serum P1NP. Risperidone was administered to female B6 mice by subcutaneously implanted osmotic minipump for four weeks, starting at 8 weeks of age. (A) Lean mass and fat mass were measured by DXA. (B) serumInsulin tolerance test. (C) Glucose tolerance test. (D) Total and femur aBMD were measured by DXA. (E) Serum P1NP and CTx were measured after an overnight fast. Bars represent mean ± standard error of untreated (white) and risperidone treated mice (black). N=5. *p<0.05 by Student’s t-test.

Figure 5

Risperidone induces osteoclastogenesis in vitro. Whole bone marrow was cultured in the presence of M-CSF and RANKL to induce osteoclastogenesis. From day 0, wells were also treated with 0, 2.5 or 25 ng/ml risperidone with each media change. Cells were fixed on day 6 and stained for TRAP. Osteoclasts (positive for TRAP and containing 3 or more nuclei) were counted. Bars represent mean ± standard error. N=3. *p<0.05 by Student’s t-test.