Blood plasma and serum contain factors that activate inwardly rectifying GIRK1/GIRK4 K+ channels in atrial myocytes via one or more non-atropine-sensitive receptors coupled to pertussis-toxin-sensitive G-proteins. This channel is also the target of muscarinic M(2) receptors activated by the physiological release of acetylcholine from parasympathetic nerve endings. By using a combination of HPLC and TLC techniques with matrix-assisted laser desorption ionization-time-of-flight MS, we purified and identified sphingosine 1-phosphate (SPP) and sphingosylphosphocholine (SPC) as the plasma and serum factors responsible for activating the inwardly rectifying K+ channel (I(K)). With the use of MS the concentration of SPC was estimated at 50 nM in plasma and 130 nM in serum; those concentrations exceeded the 1.5 nM EC(50) measured in guinea-pig atrial myocytes. With the use of reverse-transcriptase-mediated PCR and/or Western blot analysis, we detected Edg1, Edg3, Edg5 and Edg8 as well as OGR1 sphingolipid receptor transcripts and/or proteins. In perfused guinea-pig hearts, SPC exerted a negative chronotropic effect with a threshold concentration of 1 microM. SPC was completely removed after perfusion through the coronary circulation at a concentration of 10 microM. On the basis of their constitutive presence in plasma, the expression of specific receptors, and a mechanism of ligand inactivation, we propose that SPP and SPC might have a physiologically relevant role in the regulation of the heart.
Blood plasma and serum contain factors that activate inwardly rectifying GIRK1/GIRK4 K+ channels in atrial myocytes via one or more non-atropine-sensitive receptors coupled to pertussis-toxin-sensitive G-proteins. This channel is also the target of muscarinic M2 receptors activated by the physiological release of acetylcholine from parasympathetic nerve endings. By using a combination of HPLC and TLC techniques with matrix-assisted laser desorption ionization–time-of-flight MS, we purified and identified sphingosine 1-phosphate (SPP) and sphingosylphosphocholine (SPC) as the plasma and serum factors responsible for activating the inwardly rectifying K+ channel (IK). With the use of MS the concentration of SPC was estimated at 50nM in plasma and 130nM in serum; those concentrations exceeded the 1.5nM EC50 measured in guinea-pig atrial myocytes. With the use of reverse-transcriptase-mediated PCR and/or Western blot analysis, we detected Edg1, Edg3, Edg5 and Edg8 as well as OGR1 sphingolipid receptor transcripts and/or proteins. In perfused guinea-pig hearts, SPC exerted a negative chronotropic effect with a threshold concentration of 1µM. SPC was completely removed after perfusion through the coronary circulation at a concentration of 10µM. On the basis of their constitutive presence in plasma, the expression of specific receptors, and a mechanism of ligand inactivation, we propose that SPP and SPC might have a physiologically relevant role in the regulation of the heart.
Although the emergency physician often treats patients with multiple injuries, there are relatively few clinically relevant models that mimic these situations. To describe the changes after a hemorrhagic insult superimposed on traumatic brain injury (TBI), anesthetized and ventilated juvenile pigs were assigned to 35% hemorrhage (35H), TBI (via fluid percussion); TBI + 35H, and TBI + 40H (40% hemorrhage). Animals were resuscitated with shed blood and crystalloid. Hemodynamic, metabolic, behavioral, and histologic parameters were assessed for 48 h. In TBI, mean arterial pressure (MAP) was not significantly different from baseline. For TBI + 40H, MAP fell by 60% (p < 0.05). This was corrected with resuscitation. Interestingly, TBI + 35H did not show a fall in MAP, while in 35H, MAP was reduced similarly to the TBI + 40H group. ICP was elevated only initially in the TBI group. In TBI + 40H and TBI + 35H, ICP increased markedly with resuscitation, remaining elevated for 60 min. ICP remained at baseline with 35 H. Hemorrhagic focal cerebal contusions at the gray-white interface were observed in 3/5 of TBI + 40H and 5/7 of TBI + 35H. Despite the presence of subarachnoid hemorrhage (SAH) in all the animals in the TBI alone group, none of these animals demonstrated grossly discernible intraparenchymal injury. There was no evidence of intracranial injury in the 35H group. Only in animals receiving a secondary insult of hemorrhage following the primary TBI were cerebral contusions found. These experiments demonstrate the evolution of cerebral contusions as a form of secondary neurologic injury following resuscitation from traumatic brain injury and hemorrhage, even in the absence of significant blood pressure changes.
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