Hypothesis / aims of study
Mechanical or chemical stimuli to the bladder is transferred via bladder primary afferent nerves to the spinal cord. The neurons of the spinal cord projected from these primary afferent nerves are considered to be involved in processing input from the bladder. Medial dorsal horn, dorsal commissure, and sacral parasympathetic nucleus of the spinal cord have been reported to have projections from the bladder primary afferent nerves in rats and cats. However, little has been known in mice. Recently, increasing studies have used transgenic mice and revealed important knowledge regarding urinary dysfunction. Therefore, identification of the mouse neural control system for voiding reflex is now urgently needed. The aims of this study are to identify the bladder primary afferent pathways to the spinal cord in mice and to compare that pathways with those of rats and cats.
Study design, materials and methods
The male C57BL/6J mice were used. In study 1, neural tracing study, the bidirectional neuron tracer Cholera toxin B was injected into bladder wall. One week later, the spinal cord was dissected and the distribution of Cholera toxin B in the L6-S1 spinal cord was analyzed immunohistochemically. In study 2, spinal c-fos induction study, in continuous cystometry under urethane anesthesia, saline 1.5 ml/h (control group), saline 12.0 ml/h (fast infusion group), 1 mg/ml lipopolysaccharide 1.5 ml/h (LPS group), 50 mM adenosine triphosphate 1.5 ml/h (ATP group), and 0.5% acetic acid 1.5 ml/h (AA group) were continuously instilled into the bladder for 2 hours. The c-Fos (+) cells in the L6-S1 spinal cord were counted in four spinal cord regions: medial dorsal horn, lateral dorsal horn, dorsal commissure, and sacral parasympathetic nucleus.
Study 1, neural tracing study; Cholera toxin B positive nerve fibers were distributed to lateral dorsal horn, sacral parasympathetic nucleus, and dorsal commissure, but not to medial dorsal horn (figure 1). These findings suggested that afferent pathways from the bladder project to the spinal neurons of lateral dorsal horn, sacral parasympathetic nucleus and dorsal commissure in mice. Study 2, c-fos induction study; The count of c-Fos (+) cells/ 10 sections was 11.3 ± 2.3, 17.5 ± 6.2, 12.6 ± 1.9, 12.8 ± 5.2, and 15.3 ± 1.2 in medial dorsal horn; 14 ± 2.2, 17.5 ± 3.5, 15.8 ± 2.0, 8.6 ± 1.2, and 6.7 ± 0.7 in lateral dorsal horn; 78 ± 22, 219 ± 26, 203 ± 21, 318 ± 24, and 155 ± 1.7 in dorsal commissure; 26 ± 5.6, 96 ± 24, 89 ± 19, 90 ± 7.5, and 69 ± 11 in sacral parasympathetic nucleus (control vs. fast infusion vs. LPS vs. ATP vs. AA group, respectively, means ± SEM). Both mechanical stimuli by fast infusion and chemical stimuli by LPS, ATP, and AA instillation were revealed to induce the c-Fos (+) cells in dorsal commissure and sacral parasympathetic nucleus of the L6-S1 spinal cord (figure 2).
Interpretation of results
These findings suggest that both mechanical and chemical stimuli conveyed from the bladder are processed by the spinal neurons of dorsal commissure and sacral parasympathetic nucleus but not of medial dorsal horn and lateral dorsal horn in mice. This is not consistent with that of rats and cats, because noxious stimuli has been reported to increase the number of c-Fos (+) cells in medial dorsal horn, sacral parasympathetic nucleus, and dorsal commissure in these animals. Therefore, the neural control system for voiding reflex of mice must be different from that of rats and cats.