Robert A. Mitchell

The innervation of glomus cells, ganglion cells and blood vessels in the rat carotid body: A quantitative ultrastructural analysis

SummaryWe studied the ultrastructure of the rat carotid body and found that glomus cells (Type I cells) are of two types (A and B) based on the size of their dense-cored vesicles. Dense-cored vesicles in type A cells have a mean diameter nearly 30% larger than those in type B cells. Although we seldom found nerve endings on type B cells, at least two types of nerves end on type A cells. Axonal degeneration studies showed that more than 95% of these nerves are afferent axons which leave the carotid body in the carotid sinus nerve and have their cell bodies in the sensory (petrosal) ganglion of the glossopharyngeal nerve. Less than 5% are preganglionic efferent axons from the cervical sympathetic trunk which enter the carotid body with axons from the superior cervical sympathetic ganglion. We found no efferent axons from the glossopharyngeal nerve which end on glomus cells, although some do end on ganglion cells.Afferent and efferent nerve endings can be distinguished morphologically, although both types contain many synaptic vesicles and few large dense-cored vesicles. Synaptic vesicles in afferent nerve endings are 15% larger but 60% less numerous than those in efferent nerve endings. Large densecored vesicles in afferent nerve endings are similar in size but 80% less numerous than those in efferent nerve endings. Some regions of afferent nerve endings are presynaptic to glomus cells, some are postsynaptic, and some form reciprocal synapses. Efferent nerve endings are presynaptic to glomus cells but not in synaptic contact with afferent nerve endings.Blood vessels in the carotid body have both a parasympathetic and a sympathetic innervation. Most parasympathetic vasomotor nerves arise within the carotid body from ganglion cells whose preganglionic innervation is from the glossopharyngeal nerve. Terminals of these vasomotor nerves contain clear-cored synaptic vesicles. Sympathetic vasomotor nerves, most of which come from ganglion cells in the superior cervical ganglion (and from a few ganglion cells in the carotid body) have dense-cored synaptic vesicles.We postulate that (I) afferent nerve endings, which are interconnected with glomus cells by reciprocal synapses, are chemoreceptors; (2) glomus cells are dopaminergic interneurons which modulate the sensitivity of chemoreceptive nerve endings; (3) glomus cells and afferent nerves interact through reciprocal synapses which form an inhibitory feedback loop: sensory nerves release an excitatory transmitter when stimulated, the transmitter causes glomus cells to release dopamine, and dopamine inhibits the sensory nerves; (4) the feedback loop may contribute to the hyperbolic nature of the curve described by the relationship between arterial oxygen pressure and the rate of chemo-receptor firing; (5) by enhancing dopamine release from some glomus cells, preganglionic sympathetic nerves decrease chemoreceptor activity, an effect opposite from that of vasoconstriction produced by postganglionic sympathetic nerves on blood vessels j (6) synaptic interconnections enable glomus cells to influence one another. We cannot exclude the possibility that glomus cells, like afferent nerve endings, are chemoreceptors sensitive to hypoxia and hypercapnia or that glomus cells, in addition to their other functions, secrete a polypeptide hormone.

Neurogenic inflammation in the rat trachea. II. Identity and distribution of nerves mediating the increase in vascular permeability

SummaryThis study addresses the question of whether increased vascular permeability, which is a prominent feature of neurogenic inflammation in the respiratory tract, is mediated by sensory axons that end near venules in the airway mucosa. In these experiments, neurogenic inflammation was produced in the tracheal and bronchial mucosa of atropine-treated Long-Evans rats by electrical stimulation of the left or right superior laryngeal nerve and/or cervical vagus nerve. The particulate tracer Monastral blue was injected intravenously to localize the sites of increased vascular permeability, and microspectrophotometry was used to measure the amount of extravasated Monastral blue in the trachea and thereby quantify the increase in vascular permeability. In some rats, selective denervations were made to locate the cell bodies of neurons that mediate the increase in vascular permeability; in others, fluorescence immunohistochemistry and quantitative electron microscopic methods were used to determine which structures in the tracheal mucosa are innervated by these neurons. The study revealed that the vagally mediated increase in vascular permeability was sudden, transient (half-life=2.4 min) and restricted to venules. Stimulation of the left or right superior laryngeal nerve increased the permeability of venules in the extrathoracic trachea, whereas stimulation of either vagus nerve increased vascular permeability in the intrathoracic trachea and bronchi. All nerves had bilateral effects in the trachea, but the vagus nerves had largely unilateral effects in the bronchi. Neurons that mediated the increase in venular permeability had their cell bodies in the jugular (superior sensory) ganglion of the vagus nerve or rostral portion of the nodose (inferior sensory) ganglion. Preganglionic autonomic vagal neurons in the brain stem were not essential for this increase in venular permeability. Few nerves identifiable by substance P-immunohistochemistry or electron microscopy were located near the affected venules, and no nerves were within 1 μm of the walls of venules. However, the epithelium and arterioles of the airway mucosa were densely innervated. All intraepithelial nerves were within 0.1 μm of epithelial cells, and at least two-thirds of nerves near arterioles were within 1 μm of the vessel walls. We conclude that the increase in venular permeability associated with neurogenic inflammation in the trachea and bronchi of rats is mediated by sensory axons that travel in the vagus nerves and superior laryngeal nerves. We question whether tachykinins from the sensory nerves mediate the increase in vascular permeability through a direct action on venules, and raise the possibility that these nerves evoke the release from epithelial cells of mediators that contribute to the increase in vascular permeability.

Effects of dopamine, norepinephrine and 5-hydroxytryptamine on the carotid body of the dog☆

The effects of bolus intracarotid (IC) infusions of dopamine (DA), norepinephrine (NE), and 5-hydroxytryptamine (5-HT) on activity in single or few-fiber carotid chemoreceptor afferent nerve preparations were studied in pentobarbital anesthetized dogs. In addition, the effects of intravenous (IV) infusions of DA were also assessed. IC injections of DA (10 microgram) and (5-HT) (1 microgram) consistently produced a burst of intense activity followed by a period of inhibition. A similar effect was seen with IC NE (20--40 microgram) injections, but the burst of excitation occurred in only 45% of the injections. Inhibition of activity was seen in 88% of the IC NE injections. Low IC doses of all 3 amines produced inhibition of chemoreceptor afferent activity. High doses of DA IV (approximately 60 microgram/kg/))produced excitation followed by depression, while lower doses (approximately 21 microgram/kg) produced only inhibition. The excitatory effects of all 3 amines were blocked by d-tubocurarine (50--435 microgram/kg IV). Inhibitory effects of all 3 amines were blocked by dihydroergotamine (140--270 microgram/kg). The inhibitory effect produced by DA was specifically blocked by haloperidol (50--400 microgram/kg IV). We conclude that DA, NE and 5-HT can modulate carotid body activity by increasing or decreasing responses to physiologic stimuli.

Power spectral analysis of inspiratory nerve activity in the decerebrate cat

To investigate the high frequency oscillations observed in the inspiratory activity of respiratory motor nerves of decerebrate cats, we applied a signal processing technique, power spectral analysis, to the electrical activity of the phrenic and recurrent laryngeal nerves. We found two peaks in the phrenic nerve power spectral densities, one at 88.1 +/- 6.4 Hz (mean +/- S.D.) and the other at 37.1 +/- 9.7 Hz, and two peaks for the recurrent laryngeal nerve, at 87.4 +/- 10.1 Hz and at 55.4 +/- 5.1 Hz. We identified 3 factors affecting the peaks. Anesthetics reduced or eliminated the 88 Hz peak and produced new low frequency peaks in the phrenic and recurrent laryngeal nerves. Increasing end-tidal CO2 decreased the bandwidth of the 88 Hz peak and increased its amplitude relative to that of the low frequency peak. Decreasing body temperature from 38 to 30 degrees C reduced the frequency of the 88 Hz peak by 5.0 Hz/degrees C. The power spectral density of the phrenic nerve activity differed from that of the recurrent laryngeal nerve activity because the single fibers in each nerve had different power spectral densities. About 70% of the fibers recorded in a nerve had power spectral densities similar to that of the whole nerve. A minority of the phrenic nerve fibers had the same low spectral peak as the recurrent laryngeal nerve, and conversely, a minority of the recurrent laryngeal fibers had the same low spectral peak as the phrenic nerve. Bilateral removal of the dorsal respiratory group eliminated the high frequency peak in the power spectral density of the phrenic nerve and the peripheral reflexes, but rhythmic bursts of inspiratory activity remained. From these findings we hypothesized that there are two central respiratory pattern generators in the brain stem with parallel pathways to the respiratory motoneurons.

Potencies of doxapram and hypoxia in stimulating carotid-body chemoreceptors and ventilation in anesthetized cats.

The effects of doxapram on carotid chemoreceptor activity and on ventilation (phrenic-nerve activity) were tested before and after denervation of the peripheral chemoreceptors in cats. Doxapram was found to be a potent stimulus to the carotid chemoreceptors; the stimulation produced by 1.0 mg/kg doxapram, iv, equalled that produced by a Pao2 of 38 torr. Doxapram also increased phrenic-nerve activity in doses as low as 0.2 mg/kg, iv. After denervation of the peripheral chemoreceptors, doxapram in doses as large as 6 mg/kg failed to stimulate ventilation. It is concluded that (in anesthetized cats) doxapram in doses of less than 6 mg/kg increases ventilation by direct stimulation of the carotid, and, probably, the aortic, chemoreceptors, not by a direct effect on the medullary respiratory center.

In vivo activity of tracheal parasympathetic ganglion cells innervating tracheal smooth muscle

In vivo intracellular recording and intrasomal injection of Lucifer yellow revealed two populations of postganglionic parasympathetic neurons in the tracheal ganglia of cats. One consisted of large cells that had an inspiratory rhythm, had a significant post-spike afterhyperpolarization, and projected to the tracheal smooth muscle. The second consisted of small cells that fired with an expiratory rhythm, had no significant afterhyperpolarization, and projected to the intercartilaginous spaces.

Neural regulation of respiration.

We would suggest that during the evolution of the mammalian respiratory neural networks the primitive centers in the cervical cord as well as the ventral respiratory group which evolved in fish have been preserved and are capable of functioning in the absence of the dorsal respiratory group generator which evolved with air breathing. We believe that these pattern generators are separate from the identified respiratory units that have so far been studied and that the apparent reciprocal inhibition observed in the identified cells results from synchronized excitatory and inhibitory inputs arising from the pattern generator itself. We believe that the model of such a system shown in Figure 3 is consistent with the observations cited in the previous section and inconsistent with models involving a single site for a pattern generator or interaction between various populations of known or identified respiratory units.

Properties of apneusis produced by reversible cold block of the rostral pons.

Reversible cold block of the rostral pons was used to compare properties of normal and apneustic respiration in anesthetized, vagotomized, artificially ventilated cats. During apneusis we observed high frequency oscillations (HFO) in phrenic nerve activity which were reduced in frequency compared with those during a normal inspiration. Apneusis produced by mid-pontine transection or punctate pneumotaxic center (PC) lesion produced similar HFO changes. The minimal intensity of superior laryngeal nerve electrical stimulation needed to terminate a breath was higher early in an apneusis than at the same time during a normal breath. Later in apneusis the intensity required became constant and was approximately the same as that needed to end a normal inspiration at its natural termination. With intact vagi lung inflation produced a greater prolongation of expiration during apneustic respiration than during normal respiration. Apneustic type activity was observed in both phrenic and vagal inspiratory motoneurons. We suggest that: (1) HFO are generated without the PC, but the PC elevates the oscillation frequency; and (2) apneusis may result in part from a delayed activation of the normal inspiratory off-switch mechanism.

Mechanism of Cardiogenic Shock

The acute hemodynamic changes that result from coronary embolization were studied in the intact dog. Coronary embolization with lycopodium spore suspension resulted in an immediate and marked decrease in cardiac output, hypotension to shock level, elevation of pulmonary arterial and left atrial pressures, and marked increase in total peripheral resistance. These profound hemodynamic responses were partially blocked in dogs previously atropinized. The therapeutic implications of the results were discussed.

Mechanism of the isoproterenol hyperpnea in the cat

To clarify the role of peripheral chemoreceptors in the abrupt hyperpnea induced by isoproterenol injection, we measured, in anesthetized cats, the time course of VE, PETCO2, H.R. and B.P. following i.v. bolus injection of 0.5--2 microgram isoproterenol before and after bilateral section of the carotid sinus (csx), aortic (ax) and vagus (vx) nerves. We compared the hyperpneic response of isoproterenol to that of 100 microgram injections of NaCN (CN), a drug known to stimulate peripheral chemoreceptors, during air and 100% O2 breathing. The ventilatory response to isoproterenol persisted for over 90 s, whereas the CN response lasted only 30 s. Also 100% O2 markedly attenuated the CN hyperpnea but had little effect on the ventilatory response to isoproterenol. The maximum increase in ventilation in response to isoproterenol was reduced by approximately 1/3 by csx, 1/2 by combined csx and ax, and 2/3 by combined csx, ax and vx. The residual hyperpnea after csx, ax, and vs is delayed in time and lagged behind the increase in PETCO2. It is concluded that the peripheral chemoreceptors and possibly vagal afferents play a major role in the hyperpnea caused by isoproterenol, but in their absence central chemoreceptors respond to the increased PaCO2 induced by the elevated cardiac output to stimulate ventilation.