Possible role of substance P in the ischemia-reperfusion injury in the isolated rabbit lung.

The origin of the endothelial damage leading to the ischemia-reperfusion injury after lung transplantation has not been elucidated. We postulated that neurotransmitters released during the preservation of the donor lung might explain this vascular derangement. Thus, in isolated rabbit lungs preserved over 24 hours, we evaluated the release of acetylcholine (ACh) and substance P (SP), the activity of their major degrading enzymes, acetylcholinesterase (AChE) and neutral endopeptidase (NEP), and changes in the capillary permeability. Both neurotransmitters showed the highest release rate in the first 15 minutes, followed by a sharp exponential decrement at 1, 6, 12 and 24 hours. AChE and NEP activities showed no variation at these time intervals. Basal capillary permeability significantly increased (P<0.01) after 24 hours preservation with saline. This increased permeability was avoided (P<0.01) by the SP fragment 4-11 (an SP receptors antagonist), but not by atropine. These results suggest for the first time a pathogenic role of SP in the ischemia-reperfusion injury, and thus the potential usefulness of SP antagonists as additives in the lung preservation solutions should be explored.

Acetylcholine and tachykinins involvement in the caffeine-induced biphasic change in intracellular Ca2+ in bovine airway smooth muscle.

1. Caffeine has been widely used as a pharmacological tool to evaluate Ca(2+) release from the sarcoplasmic reticulum in isolated smooth muscle cells. However, in nervous tissue this drug also causes neurotransmitters release, which might cause additional effects when smooth muscle strips are evaluated. To assess this last possibility, simultaneous measurements of contraction and cytosolic Ca(2+) concentration (using Fura-2/AM) were carried out in bovine airway smooth muscle strips during caffeine stimulation. 2. A first stimulation (S1, n=11) with caffeine (10 mM) induced a biphasic change in cytosolic Ca(2+), which consisted of a transient Ca(2+) peak (254+/-40 nM, X+/-SEM) followed by a plateau (92+/-13 nM), and a transient contraction (204.72+/-31.56 mg tension mg tissue(-1)). A second caffeine stimulation (S2) produced a similar response but these parameters had a different magnitude. The S2/S1 ratios for these parameters were 0.69+/-0.02, 0.83+/-0.06 and 1.01+/-0.03, respectively. Addition of omega-conotoxin GVIA (1 micro M) and tetrodotoxin (3.1 micro M) before S2 significantly diminished these S2/S1 ratios (0.26+/-0.05, 0.26+/-0.09 and 0.64+/-0.11, respectively, n=5, P<0.05), implicating the neurotransmitters release involvement in the response to caffeine. A similar effect (P<0.01) was observed with atropine (1 micro M, n=4), the fragment 4-11 of substance P (SP) (an SP receptor antagonist, 10 micro M, n=5), and with both substances (n=4). 3. We discarded a direct effect of omega-conotoxin GVIA (1 micro M) plus tetrodotoxin (3.1 micro M) or of atropine (1 micro M) plus SP fragment 4-11 on smooth muscle cells because they did not modify caffeine responses in isolated tracheal myocytes. 4. We confirmed by HPLC that caffeine increased the release of acetylcholine (from 0.43+/-0.19 to 2.07+/-0.56 nM mg tissue(-1), P<0.02) in bovine airway smooth muscle strips. Detection of substance P by ELISA was not statistically different after caffeine stimulation (geometric means before and after caffeine, 0.69 vs. 1.97 pg ml(-1) mg tissue(-1), respectively, P=0.053). 5. We concluded that acetylcholine and tachykinins release are involved in the caffeine-induced biphasic changes in cytosolic Ca(2+) concentration.