Biomedical subjects
Publications and source records attributed to R M Harper.
We describe a low cost system for acquiring electrophysiological signals during magnetic resonance imaging. The system consists of high common-mode-rejection and low noise operational amplifiers, coupled by fiber optic cables to a receiver located at the periphery of the magnetic field. The system minimizes noise introduction which would contaminate image signals.
The objective was to determine ventral medullary surface responses to blood loss sufficient to induce shock. We examined changes in scattered light from rostral and intermediate areas of the ventral medullary surface in four intact, drug-free cats during acute hypovolemia. Scattered light images, collected during 660 and 560 nm illumination to measure cellular activity and hemodynamic aspects, respectively, were digitized at 50 frames/s during baseline, and during withdrawal of 20-30% blood volume. Hypovolemia elicited a profound hypotension and eventual bradycardia. In all cats, a modest increase in ventral medullary surface reflectance (activity decline) accompanied initial blood loss; as hypovolemia continued, and blood pressure declined, reflectance switched to a decline (activity increase), with the lowest reflectance occurring at maximal blood loss. Hypovolemia elicited multiple transient physiologic behaviors, including tachycardia, tachypnea, intermittent isolated and sustained bursts of enhanced inspiratory efforts, and extensor activation of the somatic musculature. The phasic physiological behaviors during hypovolemia were accompanied by partial recovery of medullary surface reflectance and blood pressure towards baseline values; however, reflectance continued to decrease as blood pressure progressively fell after these recovery efforts. Patterns of reflectance were not uniform over areas examined; isolated regions of enhanced or diminished reflectance appeared upon the overall images. Optical signals indicating hemodynamic changes followed the neural activity patterns, but not precisely. Regions within the ventral surface are responsive to hypovolemia, and to transient behaviors associated with momentary restoration of blood pressure; these ventral surface areas may assume essential roles in the systemic response to hypovolemic-induced shock.
Congenital central hypoventilation syndrome (CCHS) is a rare and unique condition that may prompt unparalleled approaches to the discovery of genes involved in development of cardiorespiratory control and gas exchange homeostasis. Its higher risk of recurrence in families and its association with Hirschsprung's disease suggest that an underlying genetic mechanism is involved. However, screening for mutations of the receptor tyrosine kinase RET and endothelin 3 has revealed only occasional patients affected by these mutations, therefore suggesting that CCHS may result from disruption of more than a single gene. In recent years, three principal issues have become apparent: 1) the autonomic nervous system is involved universally in CCHS cases, albeit to a varying extent; 2) the use of novel functional imaging approaches incorporating refined stimulus paradigms may provide essential research and clinical insights into localization and assessment of neural sites underlying the phenotypic expression of this syndrome; and 3) efforts to transition patients' nocturnal respiratory support to a noninvasive ventilatory modality should be critically evaluated and pursued, when appropriate, to improve the quality of life for patients and families.
We hypothesized that spontaneous activity declines over widespread areas of the cat ventral medullary surface (VMS) during rapid eye movement (REM) sleep. We assessed neural and hemodynamic activity, measured as changes in reflected 660- and 560-nm wavelength light, from the VMS during sleep and waking states in five adult, unrestrained cats and in two control cats. Relative to quiet sleep, overall activity declined, and variability, assessed by standard deviation, increased by 25% during REM sleep. Variability in activity during waking also increased by 45% over quiet sleep, but mean activity was unchanged. REM sleep onset was preceded by a reduction in the hemodynamic signal from 5 to 60 s before neural activity decline. The activity decline during REM sleep, previously noted in the goat rostral VMS, extends to intermediate VMS areas of the cat and differs from most neural sites, such as the cortex, hippocampus, and thalamus, which increase activity during REM sleep. The activity decline during REM sleep has the potential to modify VMS responsiveness to baroreceptor and chemoreceptor challenges during the REM state.
During periods of regular breathing, heart rate is slower and more variable in healthy (no history of respiratory distress syndrome) prematurely born infants with persistent apnea, relative to full-term control infants of comparable post-conceptional ages. We tested the hypothesis that the cardiovascular differences may be linked to the persistent apnea, rather than premature birth, by assessing heart rate and variability in full-term infants with persistent apnea. Thus, resting heart rate and variability were compared in full-term infants with apnea of infancy, prematurely born infants with persistent apnea, and full-term control infants. Full-term infants with persisting apnea showed slower heart rates than control infants, beginning at 4 months after birth, and enhanced heart rate variability beginning at 6 months. Healthy prematurely born infants with persistent apnea showed cardiovascular alterations similar to those of full-term infants with apnea; these alterations differed from those observed in very premature infants with histories of respiratory distress. The postnatal development of cardiovascular aberrations in infants with persistent apnea suggests that mechanisms accompanying apneic events may contribute to long-term alterations in autonomic control.
Single cell discharge, EEG activity, and optical changes accompanying alterations in breathing patterns, as well as the knowledge that respiratory musculature is heavily involved in movement and other behavioral acts, implicate hippocampal regions in some aspects of breathing control. The control is unlikely to reside in oscillatory breathing movements, because such patterns emerge in preparations retaining only the medulla (and perhaps only the spinal cord). However, momentary changes in breathing patterns induced by affect, startle, whole-body movement changes, or compensatory ventilatory changes mediated by rostral brain regions likely depend on hippocampal action in aspects of control. Hippocampal activity was enhanced prior to sighs, and this enhancement was accompanied by increased slow theta activity. Theta frequency increased during apnea, prior to return of breathing. Consideration of hippocampal contributions to breathing control should be viewed in the context that significant interactions exist between blood pressure changes and ventilation, and that modest breathing challenges, such as exposure to hypercapnia or to increased resistive loads, bring into action a vast array of brain regions involving nearly every level of the neuraxis.
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Although sessile in nature, plants are able to use a number of mechanisms to modify their morphology in response to changing environmental conditions. Differential growth is one such mechanism. Despite its importance in plant development, little is known about the molecular events regulating the establishment of differential growth. Here we report analyses of the nph4 (nonphototropic hypocotyl) mutants of Arabidopsis that suggest that the NPH4 protein plays a central role in the modulation of auxin-dependent differential growth. Results from physiological studies demonstrate that NPH4 activity is conditionally required for a number of differential growth responses, including phototropism, gravitropism, phytochrome-dependent hypocotyl curvature, apical hook maintenance, and abaxial/adaxial leaf-blade expansion. The nph4 mutants exhibited auxin resistance and severely impaired auxin-dependent gene expression, indicating that the defects associated with differential growth likely arise because of altered auxin responsiveness. Moreover, the auxin signaling events mediating phototropism are genetically correlated with the abundance of the NPH4 protein.
1. The aim of the present study was to determine the brain sites mediating aspects of respiratory and cardiovascular control in adult humans using non-invasive functional magnetic resonance (fMRI) procedures, thereby avoiding the spatial and temporal sampling limitations associated with classic neural assessment techniques. 2. We examined activity changes across the entire brain following application of respiratory loads and upon induction of blood pressure and heart rate alterations. Magnetic resonance signals were visualized with a 1.5 Tesla scanner in healthy volunteers (22-52 years of age) using procedures that optimally assess changes in brain tissue microcirculation. Images were collected during a Valsalva manoeuvre, inspiratory loading, hypercapnia, cold pressor challenges to the hand and forehead and during intervening baseline states. 3. Image values from experimental conditions were compared with corresponding baseline values on a pixel-by-pixel basis to identify brain regions in which the experimental conditions produced physiological activation. 4. Ventilatory and pressor challenges elicited significant changes in regional image signal intensity in areas within the orbital cortex, amygdala, hypothalamus and hippocampus. Cerebellar, medullary and pontine areas were also recruited. However, while particular brain regions were only activated during specific stimuli, other regional signal changes occurred with multiple experimental manipulations. 5. The findings indicate that respiratory and cardiac challenges elicit discrete activity changes over multiple brain sites. Activated regions include structures not often related to respiratory or cardiovascular regulation, such as the cerebellum; a prominent role for limbic forebrain structures in mediating the response is also suggested. The fMRI visualization procedures may greatly assist in the determination of neural structures that mediate respiratory and cardiovascular control in humans.
We developed a miniaturized, high-sensitivity camera that can be placed in areas of difficult access in freely behaving animals for neural tissue imaging. The device consists of a charged coupled device (CCD) chip, a coherent image conduit and miniature light emitting diodes (LEDs). An amplifier circuit is constructed on the camera chip and nine wires are attached for external connections. Placement of LEDs around the image conduit perimeter provides dark-field illumination, which increases detection of cellular-related light scattering changes and doubles the depth-of-view over conventional reflectance imaging procedures. The device has been successfully used to record from several deep brain structures, including the ventral medullary surface of sleeping and waking cats. The procedure allows assessment of light scattering changes that result from neural activity or detection of vital dyes to metabolic or voltage-induced activation.
At equivalent post-conceptional ages, prematurely-born infants have higher heart rates and reduced heart rate variability, relative to full-term neonates. Premature birth might exert long-lasting effects on central and peripheral mechanisms that control cardiovascular activity. We assessed development of heart rate and heart rate variability in symptomatic preterm infants up to 6 months of age. Fifty 6.5-h evening recordings of EKG and breathing were obtained from prematurely-born infants (gestational ages: 24-35 weeks). Cardiac R-R intervals were captured with a resolution of +/- 0.5 msec. One-min epochs were selected from three periods of regular respiration in recordings from premature infants and 72 recordings of full-term infants at comparable post-conceptional ages. Mean heart rate and heart rate variability were determined for each recording. At 40 weeks post-conception, prematurely-born infants with apnea of prematurity showed higher heart rates and reduced heart rate variability than did full-term neonates. These differences between premature and full-term infants persisted throughout the next 6 months in those infants born prior to 30 weeks gestation, and in those infants born at 30-35 weeks who experienced respiratory distress syndrome (RDS) during the neonatal period. The findings suggest that premature delivery, or complications thereof, exerts long-lasting effects on cardiac control.
We monitored the spatiotemporal organization of cellular activity in the medial paraventricular hypothalamus during spontaneously-occurring periods of increased inspiratory effort followed by prolonged respiratory pauses (sigh/apnea) in the freely-behaving cat. Paraventricular hypothalamic activity was assayed by video images of light captured with a stereotaxically-placed fibre optic probe. Respiratory activity was measured through electromyographic wire electrodes placed in the diaphragm. Sigh/apnea events appeared in all behavioural states, and especially during quiet sleep. Overall paraventricular hypothalamic activity declined transiently, with the onset of decline coinciding with the beginning of the sigh inspiratory effort, reached a nadir at apnea onset 4.4+0.5 s from the beginning of the sigh, increased during the course of the apnea, and subsequently rebounded above baseline to peak at 10.9+2.5 s after sigh onset. Scattered, small areas of the imaged region were activated or depressed independently of the overall image values. The data suggest that paraventricular hypothalamic activity changes dynamically during phasic respiratory events, and may contribute to the progression of the sigh/apnea. We speculate that the medial paraventricular hypothalamus influences breathing patterns through projections to parabrachial respiratory phase-shift regions, and that longer-latency influences may also be exerted indirectly through blood pressure effects from paraventricular hypothalamic projections to medullary cardiovascular nuclei. Additionally, the paraventricular hypothalamus may convey respiratory influences from other rostral structures, such as the hippocampus.
Hypothalamic warming induces panting in cats; in young kittens, panting is interspersed with slower breathing periods. The nature of neural mechanisms underlying these interspersed periods of nonpanting polypnea is unclear. We determined developmental characteristics of nonpanting breathing during thermal stress in kittens and adult cats. Warming electrodes were surgically placed into the preoptic/anterior hypothalamus (POAH) of 19 kittens and six adult cats. After surgical recovery, the hypothalamus was warmed bilaterally during sleep in unrestrained animals. Epochs of nonpanting breathing during periods of POAH warming within quiet sleep (QS) and rapid eye movement sleep (REM) were compared to control periods. Nonpanting polypnea within QS showed inspiratory and expiratory time reductions in the same proportions as those of panting. During REM, increased breathing rates accompanied POAH warming, but inspiratory/expiratory timing characteristics did not differ significantly from those of baseline breathing, except in the youngest kittens. Breathing responses to POAH warming during QS showed significant change as the kittens matured; however, response characteristics during REM did not change significantly from 12 days to adulthood. Responses to thermal stress during QS appear to depend on maturation of neural mechanisms, while separate mechanisms appear to underlie the slight respiratory rate changes during REM.
Adults with obstructive sleep apnea syndrome (OSAS) display substantial heart rate changes associated with obstructive events, and recent reports suggest similar heart rate changes in children with OSAS. These rate changes could assist screening of young patients for OSAS. Six-hour polysomnographic recordings were obtained from seven children with OSAS (mean age: 4.5 years; apnea index: 19.5 +/- 5.1) and from seven primary snorers without OSAS who served as controls (mean age: 4.7; apnea index: 0). Scatterplots of each cardiac R-R interval against the preceding interval (Poincaré plots) were used to assess beat-to-beat cardiac variability at different heart rates. Beat-to-beat variation at slow rates was significantly increased in children with OSAS relative to controls, while variation at fast and intermediate heart rates was significantly reduced in these children. We conclude that OSAS alters beat-to-beat variation in characteristic fashions in children, that the variability changes occur at all heart rates but are most significant at slow heart rates, and that these heart rate patterns could assist in screening of suspected cases of OSAS.
We assessed relationships of evoked electrical and light scattering changes from cat dorsal hippocampus following Schaeffer collateral stimulation. Under anesthesia, eight stimulating electrodes were placed in the left hippocampal CA field and an optic probe, coupled to a photodiode or a charge-coupled device camera to detect scattered light changes, was lowered to the contralateral dorsal hippocampal surface. Light at 660 +/- 10 (SE) nm illuminated the tissue through optic fibers surrounding the optic probe. An attached bipolar electrode recorded evoked right hippocampal commissural potentials. Electrode recordings and photodiode output were simultaneously acquired at 2.4 kHz during single biphasic pulse stimuli 0.5 ms in duration with 0.1-Hz intervals. Camera images were digitized at 100 Hz. An average of 150 responses was calculated for each of six stimulating current levels. Stimuli elicited a complex population synaptic potential that lasted 100-200 ms depending on stimulus intensity and electrode position. Light scattering changes peaked 20 ms after stimuli and occurred simultaneously with population spikes. A long-lasting light scattering component peaked 100-500 ms after the stimulus, concurrently with larger population postsynaptic potentials. Optical signals occurred over a time course similar to that for electrical signals and increased with larger stimulation amplitude to a maximum, then decreased with further increases in stimulation current. Camera images revealed a topographic response pattern that paralleled the photodiode measurements and depended on stimulation electrode position. Light scattering changes accompanied fast electrical responses, occurred too rapidly for perfusion, and showed a stimulus intensity relationship not consistent with glial changes.
Activity within the cat paraventricular hypothalamus (PVH) during sleep and waking states was measured by quantifying intrinsic tissue reflectivity. A fiber optic probe consisting of a 1.0 mm coherent image conduit, surrounded by plastic fibers which conducted 660 nm source light, was attached to a charge-coupled device camera, and positioned over the PVH in five cats. Electrodes for assessing state variables, including electroencephalographic activity, eye movement, and somatic muscle tone were also placed. After surgical recovery, reflected light intensity was measured continuously at 2.5 Hz during spontaneously varying sleep/waking states. Sequential state transitions from active waking to quiet waking, quiet sleep and active sleep were accompanied by progressively increased levels of PVH activity. Overall activity was highest during active sleep, and decreased markedly upon awakening. Moment-to-moment activity oscillated in the 0-0.1 Hz range, especially during active sleep and active waking; this oscillation diminished during quiet sleep. Distinct sub-regions of enhanced or diminished activity emerged within the imaged area in a state-dependent manner. We conclude that PVH activity changes with behavioral state in a regionally specific manner, and that overall activity increases during quiet sleep, and is even more enhanced in active sleep. PVH activation could be expected to stimulate pituitary release of adrenocorticotropic hormone (ACTH) and affect input to autonomic regulatory sites. Since ACTH and corticotropin releasing factor elicit arousal, and since the PVH projects to other brain areas which modulate state, we speculate that the PVH plays a role in shaping characteristics of sleep/waking states.
We recorded 259 single neurons from mesial temporal lobe structures of 21 patients with complex partial seizures. Interspike intervals within clusters of action potentials (clustered interspike intervals) recorded from cells in mesial temporal structures ipsilateral to seizure initiation were compared to clustered interspike intervals in the contralateral temporal lobe. 'Clusters' were defined as any group of three or more spikes separated by intervals of less than a defined maximum, or two spikes separated by less than half that maximum. The maximum interspike interval which defined a cluster was varied from 5 to 40 ms in 5-ms steps. Significantly smaller proportions of clustered spikes were discharged by neurons in the amygdala, hippocampus and entorhinal cortex from the temporal lobe commonly initiating seizures, compared to neurons in contralateral homotopic regions. When data from the same three structures were combined, significantly fewer cluster interspike intervals between 10 and 25 ms were recorded from cells on the side of seizure onset. Because clustered action potential discharge is a normal pattern of firing for cells that discharge endogenous bursts, the relative decrease in proportions of 10-25 ms clustered interspike intervals occurring in the temporal lobe initiating seizures might reflect a reduction in endogenous burst discharges from that side. Reduced endogenous bursting could be due to the loss of burst discharging neurons as a product of seizure-related excitotoxicity. The identification of decreased interictal single neuronal burst discharge in epileptogenic structures stresses the difference between the interictal and ictal states in patients with complex partial seizures, and the importance of the transition between those states.
Obstructive lung disease is the most common form of respiratory disturbance. However, the location of brain structures underlying the ventilatory response to resistive expiratory loads is unknown in humans. To study this issue, midsagittal magnetic resonance images were acquired in eight healthy volunteers before and after application of a moderate resistive expiratory load (30 cmH2O/liter/s), using functional magnetic resonance imaging (fMRI) strategies (1.5-T magnetic resonance; repetition time: 72 ms; echo time: 45 ms; flip angle: 30 degrees; field of view: 26 cm; slice thickness: 5 mm; 128 x 256 x 1 number of excitations). Digital image subtractions and region of interest analyses revealed significant increases in fMRI signal intensity in discrete areas of the ventral medulla, ventral and dorsal pontomedullary structures, basal forebrain, and cerebellum. Upon load withdrawal, a rapid fMRI signal off-transient occurred in all activated sites. Application of an identical load immediately after recovery from the initial stimulus resulted in smaller signal increases (P < 0.02). Prolongation of load duration was associated with progressive fMRI signal decrease across activated regions. In three additional subjects, the threshold for significant MRI signal increases was established at expiratory loads > or = 15 cmH2O/liter/s and was dose dependent with increasing loads. We conclude that resistive expiratory loads > or = 15 cmH2O/liter/s elicit regional activation of discrete brain locations in humans.