Hypothesis / aims of study
It is generally accepted that A and C fibres in the bladder pelvic nerves are the main afferents responsible for bladder sensations and micturition. In normal cats and humans, when a certain volume is reached in the bladder, the micturition is driven by the activation of bladder mechanoreceptor A delta afferents, while C afferents are sensitive to bladder cooling, over-dis ten sion, inflammation and various chemical irritants. In some pathological conditions, e.g. after spinal injury, some C afferents may be abnormally activated, thereby causing bladder over-activity.
Different from the cat, experiments in the rat indicate that some bladder C afferents are also sensitive to weak mechanical stimuli and activated by normal reflex bladder contractions. Thus, such C afferents may play the same role for bladder activities as A afferents in larger mammals. The aims of our study was to further characterize bladder C afferents in the rat by using the “marking technique” and to determine their activation profile to different manipulations of the bladder.
Study design, materials and methods
A laminectomy was made of the L5 – S1 vertebrae to allow for dorsal root single unit recordings. The L6 dorsal root was cut centrally, split into thin filaments, using microsurgical techniques, and mounted onto a pair of silver hook electrodes. Single bladder afferents were activated by electrical stimulation of the bladder pelvic nerves (1 Hz, 0.5 ms) at appropriate intensity for C afferents. Individual C units were identified by their distinct threshold and fixed, long latency to electrical stimulation of the pelvic nerves. Once identified, the “marking technique” (1) was applied by a stepwise increasing in frequency of electrical stimulation, thereby imposing a controlled decrease in conduction velocity of activated C afferents. Subsequently, their response to various bladder stimuli was evaluated. Nerve length was measured at the end of the experiment. To minimize unrelated noise, evoked unit responses were averaged and subsequently analyzed off-line with a PC based system. Experimental procedures were approved by the Animal Research Ethical Committee of Linköping.
Results
Twenty-seven afferents were identified following electrical stimulation of the ipsilateral bladder pelvic nerves. The range of latency for all units was 10 to 50 ms corresponding to a conduction velocity (CV) between 8.3 and 0.8 m/s. Units conducting ≤2.5 m/s were classified as C fibres (n=18), and those conducting more rapidly as A (n=9). The C units were further characterized by activity dependent CV slowing, induced by increasing the frequency of electrical stimulation. At 3, 5 and 10 Hz, the mean CV of C units decreased by 1.7, 10.9 and 17.4 % to the control respectively. All tested C units were affected at stimulation frequencies above 5 Hz, while A units were unchanged. Some C units were responsive to high isotonic pressure manifested by a decrease in their CV at 40 cmH2O, but not at 20 cmH2O. Almost all tested C units were sensitive to the acidic solution (pH 4), potassium chloride 150 mEq/L or 0.01% menthol.
Interpretation of results
The marking technique was used in this study by recording bladder afferent activities in the L6 dorsal root following electrical stimulation of bladder pelvic nerves. All identified C afferents decreased conduction velocity at increased stimulation frequency and this change was utilized to evaluate their response to imposed increase in bladder pressure, low pH, KCl or menthol. With this technique, the activation pattern of several C units can be determined simultaneously during in vivo manipulations of the bladder.
Concluding message
By using the “marking technique”, we were characterized bladder C afferents in the rats and identified their activation profile to different manipulations of the bladder, such as imposed increase in bladder pressure, low pH, KCl or menthol etc., those abnormally activated bladder C afferents might be the causing of bladder over activity.