Bradykinin Research
BK Effects
In order to determine the effects of BK on the neurones a number of measurements were taken in control situation and then again following administration of 100nM (nanomoles) of BK. The measurements gained information about all aspects of the neurones’ electrical activity and response. These are the factors likely to be affected by BK, in particular the Resting Membrane Potential and the Threshold through sensitisation.
Control Results
- Resting Membrane Potential
- The Resting Membrane potential (Vm), the current across the membrane that is present when the cell is at rest, was measured in mV (milliVolts). The measurments ranged from –43 to –70 mV (Average = -56.25 mV). During experimentation the Vm was maintained at exactly 60mV in all cells in order to level the readings for simplified interpretation.
- Threshold
- This is the current required in order to produce an action potential in
the cell being measured. A series of graduating currents were applied to
each cell and the threshold at which an Action Potential was produced was
recorded.
The measurements ranged from –19 to –39.5 mV (Average = 27.14 mV). - Action Potential Measurements
- A number of measurements were taken from each neurone for its action potential:
- AP: Action Potential Peak Voltage (mV)
- AHP: After-Hyperpolarisation (mV)
- AP0R: Time where the rising action potential crosses the 0 mV line. (ms)
- APP: Time of the Action Potential Peak (ms)
- AP0F: Time where the falling action potential crosses the 0 mV line. (ms)
- AHPS: Start time of the AHP (ms)
- AHP80: Time for the AHP to reach 80% of recovery (ms)
- APR: Action Potential Rise Time (ms)
- APF: Action Potential Fall Time (ms)
- APL: Action Potential length (duration between points 2 & 4) (ms)
These measurements allowed the Action Potentials to be plotted and for subsequent comparisons to be made after the addition of chemicals. The results are tabulated below with the post-administration measurements.
Post-Administration
These were taken after the administration of 100nM of bradykinin (BK).
- Input Resistance
- measured in M
(Megaohms). Of the 18 neurones the input resistance ranged from 56 to 330
M,
the average being 163.69 M.
BK insignificantly affected the resistance. - Spontaneous Firing
- One of the neurones exhibited spontaneous firing following the administration of BK.
- Humps
- Out of the 18 neurones examined 15 displayed a characteristic hump. Neurone 1 developed a hump and there were no changes to neurones 5 and 11 previously found to have no hump.
- Sag
- 11 out of the 18 neurones displayed this characteristic. All neurones were unchanged except for Neurone 12, which originally displayed a hump but did not after BK administration.
- Resting Membrane Potential
- The measurments following BK administration ranged from –41 to –68 mV (Average = -51.56 mV). A small depolarisation in the natural membrane potential was observed, from –56.25 to –51.56 (ie. –56±2mV to –52±2mV). This change took place over a time delay of approximately 23 ±3 s between the application of BK and the measured response. As already mentioned, the input resistance did not alter with the administration of BK, suggesting that there was no association between the depolarisation of membrane potential and input resistance.
- Threshold
- In nearly all neurones tested the threshold dropped in response to BK, one neurone’s threshold stayed the same, and none were increased. The post-administration range was from -18mV to –43mV, with the average being –29.5mV. The margin by which the threshold fell was consistently 2mV ±1mV.
- Action Potential Measurements
- The same 10 measurements were taken from each neurone for its action potential. The comparison of these measurements with pre and post BK administration data is shown in the table below.
| Measurement | Average Pre-BK Administration | Average Post-BK Administration |
|---|---|---|
| AP (mV) | 45.83 |
45.75 |
| AHP (mV) | -61.33 |
-61.00 |
| AP0R (ms) | 5.95 |
5.97 |
| APP (ms) | 6.57 |
6.56 |
| AP0F (ms) | 7.91 |
7.90 |
| AHPS (ms) | 8.79 |
8.72 |
| AHP80 (ms) | 46.06 |
46.97 |
| APR (ms) | 0.63 |
0.63 |
| APF (ms) | 1.33 |
1.31 |
| APL (ms) | 1.96 |
1.93 |
The results indicate that the measurements taken within the Action Potential are unchanged by the administration of BK. Thus, BK does not affect the height, duration or recovery of the action potential.
- Effects of Sag & Hump
- The neurones that had a characteristic sag on the injection of a hyperpolarizing current did not appear to have any significant differences in either the original AP shape of in the response to BK. The same was true for the neurones that displayed a characteristic hump in their AP. The measurements for the different groups were compared and no significant differences were seen.
NB. All averages taken were subject to a standard error of ± 2mV unless otherwise stated.
Discussion
The results indicate that BK has no effect on the majority of measurements within the Action Potential. However, it has a marked and consistent effect on the Threshold and Membrane Potential of each neurone. Both of these factors are consistently depolarised by approximately 2mV following the administration of 100nM BK. Consequently it is clear that BK does act on CNS Trigeminal neurones to produce the same sensitisation (depolarised membrane potential & lower threshold) & activation (produces APs) that are seen in the periphery. As such its role must be further investigated in order to establish its potential involvement in the pathogenesis of migraine – this is undertaken in BK Receptors.
Graph showing sensitisation. Click the thumbnail above for a larger version.
Graph showing BK Activation. Click the thumbnail above for a larger version.
