John H. Schild - US grants

Neurons within the dorsomedial region of the nucleus of the tractus solitarius (mNTS) are essential to the baroreflex, which provides rapid dynamic regulation of the cardiovascular system within a single heartbeat. The objectives of this research are to investigate the intrinsic firing properties and signal processing characteristics of the individual neurons and synaptic interconnections which constitute this medullary cardiovascular control circuitry. Adult rat brainstem will be sectioned at an angle which preserves the mNTS, providing a maximum length of the solitary tract, and will be partially submerged in a perfusion chamber for superfusion of the slice. The research investigations will determine: 1) the effects of known Ca2+-activated K+ current (l KCa) blockers on the repetitive firing characteristics of mNTS neurons, 2) the manner in which the ongoing and evoked patterns of activity in mNTS neurons are influenced by the dynamic interaction between intracellular Ca2+ and lKCa, and 3) the effects of the relative strength of lKCa, (as altered using partial pharmacological block) upon the entrainment and phase-response properties of these spontaneously oscillating mNTS neurons. It is anticipated that these results will enhance our understanding of both the functional properties of mNTS neurons and the biophysical mechanisms contributing in the processing of baroreceptor inputs to this medullary cardiovascular circuitry. Furthermore, these results could potentially reveal new insights into the neurogenesis of such clinical issues as hypertension and cardiac dysrhythmias.

[unreadable] DESCRIPTION (provided by applicant): The arterial baroreceptor (BR) reflex plays an essential role in autonomic control of the heart. Altered discharge of BR afferents occurs with hypertension and heart failure and is therefore inextricably linked to autonomic nervous system dysfunction. BR are broadly classified as myelinated or unmyelinated afferents, each exhibiting distinct discharge patterns in response to arterial pressure changes. The micromechanical environment of the peripheral termination certainly plays a role in the process of pressure transduction. However, the operational differences in sensory coding between these two functional phenotypes may also arise from unique distributions of ion channels at critical points along the afferent pathway (e.g. arterial pressoreceptor, cell body, central synapse). Unfortunately, such a bimodal demarcation belies the continuum of physiological properties exhibited by BR and makes difficult the integration of cellular and systems level observations. For example, selective recruitment of myelinated or unmyelinated BR via electrical excitation of voltage-gated ion channels (i.e. activation independent of mechanotransduction) evokes dramatically different heart rate and blood pressure reflex responses. The ionic mechanisms that contribute to the differential sensory encoding properties of myelinated and unmyelinated BR are largely unknown. Here, we use a newly developed adult rat nerve-ganglion preparation for patch clamp study of fluorescently identified aortic baroreceptor neurons which ensures unambiguous classification of sensory modality and afferent fiber type. Preliminary data are suggestive of a differential utilization of voltage- and ligand-gated ion channels that may potentially explain some of the contrasting pressure encoding properties of myelinated and unmyelinated BR. For example, neural discharge from myelinated afferents appears less dependent upon N-type Ca2+. (ICa,N)and BK-type Ca2+-activated K+ (IKCa,BK)ion channels than activity arising from unmyelinated afferents, despite voltage clamp evidence for functional coexpression of ICa,N and IKCa,BK in both phenotypes. Such differential ionic mechanisms may underlie the disparate neural encoding properties of myelinated and unmyelinated BR and could potentially influence brainstem integration of cardiovascular afferent information if similarly represented at the presynaptic terminals. These fundamental details may lead to novel pharmacological strategies in the management of cardiovascular pathologies such as acute hypertension and dysrhythmias that are well known to involve or invoke autonomic reflexes through BR activation. [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Arterial baroreceptors (BR) are essential for reliable neural control of heart rate and blood pressure. The data quantifying the properties of these pressoreceptors and the reflexogenic consequences of BR function and dysfunction are extensive. Experimental interpretations of baroreflex dysfunction and cardiovascular pathologies such as neurally mediated syncope, dysrhythmias and hypertension are of significant clinical importance. Age related changes in arterial wall properties strongly correlate with cardiovagal baroreflex impairment, increased levels of blood pressure variability, an impaired ability to respond to acute hemodynamic challenges and increased risk of sudden cardiac death (Monahan, 2007). The BR sensor itself represents an essential functional intersection across these diverse pathologies and yet a clarifying explanation of the transduction machinery is lacking. Our working hypothesis is that the microanatomy, extracellular tissue matrix, excitable neural membrane of the BR terminal complex and regional arterial wall tissues all make functionally distinct contributions to the spatial integration and transduction of localized micromechanical forces arising from arterial pressure dynamics. Our specific aims center upon the neuromechanical properties of myelinated and unmyelinated rat aortic BR as these are accessible for both micro- and macroscopic study using three complementary methodologies: 1) confocal, electron and fluorescent microscopy in conjunction with immunohistochemical labeling of protein expression within, and the tissue constructs that circumscribe, the BR terminal ending, 2) extracellular recording of aortic BR fiber discharge in response to computer controlled pressure loading of the arterial wall and 3) synthesis of these disparate microscopy and biophysical data into comprehensive computational models of the neural and micromechanical mechanisms of mechanosensory transduction that are inaccessible for direct testing and measurement. Our preliminary results illustrate: 1) essential differences between the topological distribution of molecularly identified ion channels along the ultrastructure of BR terminals with myelinated and unmyelinated fibers, 2) a minimum complement of the ion channels expressed at the cell body of BR neurons may underlie the neurogenic mechanisms of mechanotransduction and 3) that these ionic mechanisms contribute to, but cannot entirely account for, such dynamic properties as adaptation, hysteresis and resetting of discharge threshold. This combined experimental and computational strategy is producing a more biophysical understanding of the structure-function relationships associated with BR afferents relative to the basement membrane about the terminal ending, the elastin and collagen fibers within the surrounding tissue matrix as well as the neuro-integrative processes of mechanotransduction. As we recently demonstrated (Feng et al., 2007), this integrative approach can lead to new insights concerning acute cardiovascular pathologies that are well known to invoke autonomic reflexes through activation of arterial mechanoreceptors. PUBLIC HEALTH RELEVANCE: In order for the brain to properly control the heart it must continually receive information concerning blood pressure and heart rate. This application involves experimental bioengineering research related to the arterial pressure sensors (baroreceptors) that provide this critically important information to the brain. A more detailed understanding of how these sensors normally work and adapt to short and long term changes in blood pressure, heart rate and the condition of the arteries (e.g. with aging) will help physicians better manage heart function under conditions of health and disease.

DESCRIPTION (provided by applicant): Gender differences in cardiovascular function are well known. Hormones and related receptors are critically important factors, but clinical studies are revealing potential gender differences in afferent mediated autonomic nervous system (ANS) function. Some ANS assessments of cardiovagal reflexes have proven to be sexually dimorphic. In this study we investigate hypotheses related to possible neuroanatomical, neuophysiological and biophysical differences between aortic baroreceptors (BR) in male and female rats. A gender-related bias in aortic BR fiber type, pressure encoding and neurohormonal regulation may reveal as yet unrecognized mechanisms associated with noted gender differences in integrated cardiovagal control. This proposal builds upon our previous studies quantifying the differential composition of ionic channels in myelinated and unmyelinated BR afferents and the manner in which this defines the strikingly different reflex control of heart rate and blood pressure evoked by these neuroanatomically distinct afferent pathways. The first aim examines gender-related differences in rat aortic BR afferents. Morphometric analysis of aortic BR fibers and study of fluorescently identified aortic BR neurons (ABN) gives preliminary evidence that female rats have ~50% more myelinated BR, revealing, for the first time, a functionally distinct subtype of low threshold myelinated ABN rarely present (~2%) in age-matched males. The second aim quantifies the neuromodulatory capacity of estradiol (E2) upon this unique subtype of BR afferents. Nerve recordings show that E2 can increase the pressure-dependent discharge of single myelinated aortic BR fibers from female rats. Physiological levels of E2 (0.1 - 1 nM) acting, at least in part, via membrane bound estrogen receptors can selectively increase the excitability of this ABN subtype in female rats. Interestingly, E2 had no effect upon unmyelinated ABN from either gender. The third aim determines if E2 can alter K+ ion channel function in a manner consistent with the observed increased excitability. Our preliminary data show that, unlike all myelinated ABN in males and the balance of myelinated ABN from females, this unique subset expresses BKCa channels that provide ~25% of the whole cell potassium current. We further show that E2 selectively inhibits this BKCa current, offering one potential mechanism for the E2 sensitization of myelinated BR excitability in female rats. Finally, in aim four we determine if E2 can alter monosynaptic transmission of BR afferents onto 2nd-order BR neurons in the NTS. In male rats, monosynaptic transmission of myelinated afferents to the NTS does not involve BKCa channels. In stark contrast, E2 in female rats can increase monosynaptic transmission of myelinated afferent pathways with companion studies implicating a role for BKCa channels. Gender-related differences in the neural integration of BR sensory information from the afferent terminal through to 2nd-order BR neurons in the NTS could potentially lead to novel advances in the management of cardiovascular health and disease in the female population. PUBLIC HEALTH RELEVANCE: Gender differences in cardiovascular function are well known and while sex hormones are important contributing factors, there is growing evidence for alternative mechanisms. This project aims to determine if there are fundamental neuroanatomical and neurophysiological differences in the baroreceptor afferent pathway between men and women. Evidence for a gender-related difference in baroreceptors could potentially lead to novel advances in the effective management of cardiovascular health and disease in the female population.