Steven C. Hempleman - US grants

Although avian and mammalian carotid body chemoreceptors are known to be structurally similar, and to serve similar functions in the control of ventilation, the stimulus-response characteristics of single unit avian carotid chemoreceptors are poorly documented. The proposed study will establish functional similarities and differences using single unit neural recording techniques. Static and dynamic responses of avian carotid chemoreceptors to their adequate stimuli will be defined, and their responses to substances known to affect mammalian carotid bodies--eg. oligomycin, KCN, and norepinephrine--characterized. The pattern of avian carotid body discharge will be quantitated with interspike interval histograms. The independent and interactive effects of steady levels of pH, PCO2, and PO2 on avian carotid chemoreceptors will be determined from single unit stimulus-response curves. Receptor responses to hypocapnic-hypoxic stimuli will be of special interest because the superior hypoxic tolerance of birds compared to mammals may be partly due to a stronger hypoxic drive in birds. Arterial PCO2 oscillations linked to tidal breathing affect carotid chemoreceptor discharge patterns in mammals. Since arterial PCO2 oscillations are predicted to be even larger in birds, correlations will be made between avian carotid chemoreceptor discharge pattern, and the arterial PCO2 oscillations measured wiith an intravascular pH electrode. The hypothesis that dynamic responses of carotid chemoreceptors may help couple ventilation to metabolism by detecting exercise-induced changes in arterial PCO2 oscillations will be tested. Dynamic receptor responses will be quantitated while forcing arterial PCO2 oscillations of varying shape and frequency, using artificial, unidirectional pulmonary ventilation. Unidirectional ventilation of the avian lung offers a powerful tool for controlling arterial gas tensions, allowing a systems analysis approach to the study of receptor frequency response. Sinusoidal PCO2 oscillations will be used to determine the frequency response of avian carotid chemoreceptors, and ramp oscillations with varying up and down slopes, but constant period and amplitude, will be used to determine the effect of rate of PCO2 change on receptor discharge. Similar experiments would be much more difficult in mammals. The results of this study will broaden our knowledge of avian carotid body chemoreceptor responses, and serve to guide further studies of carotid bodies in general.

The discharge patterns of carotid body chemoreceptors will be studied. An avian animal model was chosen because avian carotid body chemoreceptors are nearly identical in structure and function to mammalian carotid bodies, and because the avian lung offers the powerful experimental advantage of unidirectional ventilation (UDV) for controlling arterial blood gases. Experiments are proposed that combine (UDV), on-line blood gas measurement, single unit neural recording techniques, and computerized on-line data acquisition to test the physiological responses of carotid body chemoreceptors in ways difficult or impossible with a mammalian model. Carotid body chemoreceptors are multimodal, responding to arterial PO2, PCO2, and other stimuli. This project will determine the response of single receptors to arterial pH and blood pressure. This project will also analyze the temporal occurrence of action potentials from single chemoreceptors exposed to different static levels of stimuli cause different receptor discharge patterns. Pattern differences could be a neural encoding mechanism for carrying differential stimulus information to the central respiratory controller, and may represent fundamental differences in transduction mechanisms for O2 and CO2. Dynamic oscillations of arterial PCO2 associated with tidal breathing are hypothesized to cause a feed-forward control signal via the carotid bodies for ventilatory control during exercise. This project will use UDV-induced ramp oscillations of arterial PCO2, and PO2 to test the rate sensitivity of carotid body chemoreceptors (which would amplify the feed-forward signal in exercise). UDV-induced sinusoidal oscillations of arterial PO2 and PCO2 will be used to test the frequency response of the receptors (determining the receptor response to changes in respiratory rate), and it will test for phase differences between oscillating PO2 or PCO2 receptor response (which affects the efficacy of chemoreceptor discharge arriving at the central controller). Many aspects of the normal dynamic and static carotid body chemoreceptor sensitivity remain uncertain. It is important to study these normal physiological responses so that we can define the roles of carotid body chemoreceptors in the control of pulmonary ventilation in health and disease.

[unreadable] DESCRIPTION (provided by applicant): CO2-sensing respiratory chemoreceptors are found in diverse locations in air-breathing vertebrates, including multiple sites within the brainstem, carotid and aortic bodies, and lungs, affirming their functional importance in physiological pH/PCO2 homeostasis. We have a long-standing interest in CO2 signal transduction by an unusually responsive type of CO2 sensor -the avian intrapulmonary chemoreceptor (IPC). Past IPC research indicates they (like most respiratory chemoreceptors) probably sense H+ rather than CO2 directly. In IPC, CO2-induced acidosis inhibits action potential discharge rate, making IPC an excellent model for many mammalian central and airway chemoreceptors that share this characteristic, and a good contrast to many other mammalian respiratory chemoreceptors that are excited by CO2. The first aim of this proposal will test the intracellular pH sensing hypothesis using weak and strong acids and bases that vary in their degree of ionization and membrane permeability. Weak acids and bases should have better intracellular access and greater effects on IPC discharge and CO2 sensitivity than strong acids and bases. The second aim investigates TREK-like tandem pore domain leak channels that we recently discovered in IPC. Because TREK channels are opened by intracellular acidosis, they could be the long-sought molecular target for inhibition of IPC by intracellular acidosis/CO2. This will be tested using TREK agonists or antagonists including riluzole, polyunsaturated fatty acids, arachidonic acid, cyclopropane, nitrous oxide, xenon, and osmotic challenges. The third aim will test whether peak IPC discharge rate and the magnitude of IPC spike frequency adaptation (i.e. attributes of neural coding) scale with body size in proportion to M-1/4. Such scaling would match information delivery by IPC afferent discharge to the breathing frequency of the animals, which also scales to M-1/4. To test this, dynamic IPC discharge responses to standardized CO2 step stimuli will be quantified in very small animals (~12 g body mass) and in the very large animals (~100,000 g body mass). This research will enhance understanding of respiratory chemoreceptors in general and may reveal drugs useful for treating CO2 insensitivity during respiratory failure. This research investigates fundamental neural processes that detect CO2 levels in the body and send a neural signal to the brain that ultimately controls breathing. This is important for human health, because understanding the basis of CO2 chemotransduction may help develop more effective treatments and drugs for the loss of CO2 chemosensitivity that often complicates patient survival in serious cardiopulmonary disease. [unreadable] [unreadable] [unreadable]