Substance P modulates nicotinic responses of intracardiac neurons to acetylcholine in the guinea pig.

Application of substance P (SP) to intracardiac neurons of the guinea pig causes slow depolarization and increases neuronal excitability. The present study was done to determine the influence of SP on fast excitatory responses of intracardiac neurons to ACh. Intracellular recording methods were used to measure responses of intracardiac neurons in whole mount preparations of atrial ganglionated nerve plexus from guinea pig hearts. Local pressure ejection of 100 microM SP (1 s) from a glass micropipette caused slow depolarization of all neurons (n = 38) and triggered action potential generation in 47% of the cells tested. Bath application of SP (0.5-100 microM) caused a dose-dependent depolarization of intracardiac neurons but rarely evoked action potentials, even at the highest concentration. However, such treatment with SP enhanced nicotinic responses evoked by local pressure ejections of ACh (10 mM, 10- to 100-ms duration) in 77% of intracardiac neurons studied (n = 52). A significant increase in amplitude of ACh-evoked fast depolarization occurred during treatment with 0.5 microM SP (13.0 +/- 1.8 mV for control vs. 17.7 +/- 1.9 mV with SP present, n = 7, P = 0.019). At higher concentrations of SP, enhancement of the response to ACh resulted mainly in action potential generation. However, responses to ACh were attenuated by SP in 15% of the intracardiac neurons studied. This attenuation occurred primarily during exposure to 10 and 100 microM SP and was manifest as a reduction in amplitude of nicotinic fast depolarization or inhibition of ACh-evoked action potentials. These findings support the conclusion that SP could function as a neuromodulator and neurotransmitter in intracardiac ganglia of the guinea pig.

Increased ganglionic responses to substance P in hypertensive rats due to upregulation of NK(1) receptors.

Intravenous injection of substance P (SP) increases renal nerve firing and heart rate in spontaneously hypertensive rats (SHRs) and Wistar-Kyoto rats (WKYs) by stimulating sympathetic ganglia. Blood pressure is increased in SHRs but lowered in WKYs. This study assesses the role of neurokinin-1 (NK(1)) receptors in mediating the ganglion actions of SP. Rats for functional studies were anesthetized and then treated with chlorisondamine. Renal nerve, blood pressure, and heart rate responses to intravenous injection of the NK(1) receptor agonist GR-73632 were similar but less than those to equimolar doses of SP in SHRs. GR-73632 only slightly increased renal nerve firing and heart rate and lowered blood pressure in WKYs. The NK(1) receptor antagonist GR-82334 (200 nmol/kg iv) blocked the ganglionic actions of GR-73632 and the pressor response to SP in SHRs. It reduced the renal nerve and heart rate responses by 52 and 35%. This suggests that the pressor response to SP is mediated by ganglionic NK(1) receptors and that NK(1) receptors also have a prominent role in mediating the renal nerve and heart rate responses to SP. Quantitative autoradiography showed that NK(1) receptors are more abundant in the superior cervical ganglia of SHRs. RT-PCR showed increased abundance of NK(1) receptor mRNA in SHRs as well. These observations suggest that the greater ganglionic stimulation caused by SP in SHRs is due to upregulation of NK(1) receptors.

Tachykinin receptor subtypes in the isolated guinea pig heart and their role in mediating responses to neurokinin A.

Selective tachykinin agonists were used to identify cardiac and coronary responses mediated by specific tachykinin receptor subtypes in isolated, perfused guinea pig hearts. Receptor desensitization with selective agonists and blockade with selective antagonists were used to determine the role of specific subtypes in generating responses to neurokinin A (NKA). Dose-dependent cardiac and coronary effects were evoked by bolus injections of ¿Sar(9), Met(O(2))(11)substance P (¿Sar(9),Met(O(2))(11)SP), GR64349, and ¿MePhe(7)neurokinin B (¿MePhe(7)NKB) (selective agonists for NK(1), NK(2), and NK(3) receptors, respectively). Each agonist caused bradycardia, but GR64349 was most effective (34 +/- 4% decrease in heart rate with 32 nmol, n = 8). Prominent increases in ventricular contractility and perfusion pressure also occurred with 32 nmol of GR64349 (25 +/- 6 and 33 +/- 4%, respectively). ¿Sar(9), Met(O(2))(11)SP was unique in having a high potency for decreasing ventricular contractility and perfusion pressure. Bolus injections of 25 nmol of NKA decreased rate (48 +/- 2%, n = 51), increased contractility (26 +/- 2%), and had biphasic effects on perfusion pressure (24 +/- 1% decrease followed by 9.2 +/- 1.4% increase). Desensitization with GR64349 or treatment with the NK(2) antagonist SR48968 reduced the bradycardic response to NKA by greater than 75% and eliminated the positive inotropic response. The remaining bradycardia occurred through NK(3) receptors. Desensitization with ¿Sar(9),Met(O(2))(11)SP or NK(1) blockade with FK888 eliminated the coronary relaxant action of NKA and enhanced the pressor response. It is concluded that three tachykinin receptor subtypes are present in the guinea pig heart and that each contributes to the overall response evoked by NKA.

Endogenous tachykinins cause bradycardia by stimulating cholinergic neurons in the isolated guinea pig heart.

The purpose of this study was to determine if endogenous tachykinins can cause bradycardia in the isolated perfused guinea pig heart through stimulation of cholinergic neurons. Capsaicin was used to stimulate release of tachykinins and calcitonin gene-related peptide (CGRP) from cardiac afferents. A bolus injection of 100 nmol capsaicin increased heart rate by 26 +/- 7% from a baseline of 257 +/- 14 beats/min (n = 6, P < 0.01). This positive chronotropic response was converted to a minor bradycardic effect in hearts with 1 microM CGRP-(8-37) present to block CGRP receptors. The negative chronotropic response to capsaicin was markedly potentiated in another group of hearts with the further addition of 0.5 microM neostigmine to inhibit cholinesterases. In this group, capsaicin decreased heart rate by 30 +/- 10% from a baseline of 214 +/- 6 beats/min (n = 8, P < 0.05). This large bradycardic response to capsaicin was inhibited by 1) infusion of neurokinin A to desensitize tachykinin receptors or 2) treatment with 1 microM atropine to block muscarinic receptors. The latter observations implicate tachykinins and acetylcholine, respectively, as mediators of the bradycardia. These findings support the hypothesis that endogenous tachykinins could mediate axon reflexes to stimulate cholinergic neurons of the intrinsic cardiac ganglia.

Enhanced ganglionic responses to substance P in spontaneously hypertensive rats.

Intravenous injection of substance P (SP) increases blood pressure in normotensive rats by stimulating sympathetic ganglia. This study compared the effects of SP to increase renal nerve firing and blood pressure in normotensive and hypertensive rats treated with chlorisondamine. The increase in renal nerve firing was greatest in spontaneously hypertensive rats (SHR), intermediate in Wistar rats, and least in Wistar-Kyoto (WKY) rats. Blood pressure was increased more in SHR than in Wistar rats. Blood pressure was not increased in WKY rats. Responses to the ganglionic stimulant 1,1-dimethyl-4-phenylpiperazinium were the same in the three strains. These results suggest that there is a selective increase in the action of SP on sympathetic ganglia of SHR and that ganglion responsiveness to SP is correlated with its effect on blood pressure.

Actions of tachykinins within the heart and their relevance to cardiovascular disease.

Substance P and neurokinin A are tachykinins that are co-localized with calcitonin gene-related peptide (CGRP) in a unique subpopulation of cardiac afferent nerve fibers. These neurons are activated by nociceptive stimuli and exhibit both sensory and motor functions that are mediated by the tachykinins and/or CGRP. Sensory signals (e.g., cardiac pain) are transmitted by peptides released at central processes of these neurons, whereas motor functions are produced by the same peptides released from peripheral nerve processes. This review summarizes our current understanding of intracardiac actions of the tachykinins. The major targets for the tachykinins within the heart are the intrinsic cardiac ganglia and coronary arteries. Intrinsic cardiac ganglia contain cholinergic neurons that innervate the heart and coronary vasculature. Tachykinins can stimulate NK3 receptors on these neurons to increase their excitability and evoke spontaneous firing of action potentials. This action provides a mechanism whereby tachykinins can indirectly influence cardiac function and coronary tone. Tachykinins also have direct effects on coronary arteries to decrease or increase tone. Stimulation of NK1 receptors on the endothelium causes vasodilation mediated by nitric oxide. This effect is normally dominant, but NK2 receptor-mediated vasoconstriction can also occur and is augmented when NK1 receptors are blocked. It is proposed that these ganglion stimulant and vascular actions are manifest by endogenous tachykinins during myocardial ischemia.

cAMP and in vitro inotropic actions of secretin and VIP in rat papillary muscle.

Secretin and VIP stimulate cardiac adenylyl cyclase activity and exert a positive inotropic action in several mammalian species. This study examined positive inotropic activity and cAMP levels in rat papillary muscle. Isoproterenol and secretin increased contractions by 150+/-31% and 129+/-27%, respectively. VIP increased contraction by 30+/-21% only at 10 microM. Isoproterenol significantly increased cAMP levels by 82%, whereas increases by secretin (58%) and VIP (56%) were not significant. These results are consistent with reports that secretin and VIP stimulate cardiac adenylyl cyclase in the rat, but suggest that cAMP tissue levels cannot totally explain the positive inotropic responses to secretin and VIP.

Substance P evokes bradycardia by stimulation of postganglionic cholinergic neurons.

Substance P (SP) evokes bradycardia that is mediated by cholinergic neurons in experiments with isolated guinea pig hearts. This project investigates the negative chronotropic action of SP in vivo. Guinea pigs were anesthetized with urethane, vagotomized and artificially respired. Using this model, IV injection of SP (32 nmol/kg/50 microl saline) caused a brief decrease in heart rate (-30+/-3 beats/min from a baseline of 256+/-4 beats/min, n = 27) and a long-lasting decrease in blood pressure (-28+/-2 mmHg from baseline of 51+/-5 mmHg, n = 27). The negative chronotropic response to SP was attenuated by muscarinic receptor blockade with atropine (-29 +/- 9 beats/min before vs -8 +/- 2 beats/min after treatment, P = 0.0204, n = 5) and augmented by inhibition of cholinesterases with physostigmine (-23 +/- 6 beats/min before versus -74 +/- 20 beats/min after treatment, P = 0.0250, n = 5). Ganglion blockade with chlorisondamine did not diminish the negative chronotropic response to SP. In another series of experiments, animals were anesthetized with sodium pentobarbital or urethane and studied with or without vagotomy. Neither anesthetic nor vagotomy had a significant effect on the negative chronotropic response to SP (F3,24 = 1.97, P = 0.2198). Comparison of responses to 640 nmol/kg nitroprusside and 32 nmol/kg SP demonstrated that the bradycardic effect of SP occurs independent of vasodilation. These results suggest that SP can evoke bradycardia in vivo through stimulation of postganglionic cholinergic neurons.

Blastic mantle cell leukemia: a previously undescribed form.

The leukemic phase of mantle cell lymphoma (mantle cell leukemia) is defined as an absolute lymphocyte count of greater than 4,000/microliter and characterized by the presence of relatively small, slightly irregular lymphocytes in the peripheral blood. Although a variant of mantle cell lymphoma with blastic morphsology exists and has been previously well described, a blastic morphologic variant of mantle cell leukemia has not been described. We report such a case in a 74-year-old male who presented with splenic rupture and an elevated white blood cell (WBC) count. The diagnosis was based on flow cytometric immunophenotyping and the cytomorphology of the peripheral blood.

Composite B-cell and T-cell lymphoma arising 24 years after nodular lymphocyte predominant Hodgkin's disease.

Twenty-four years after apparently successful treatment for nodular lymphocyte predominant Hodgkin's disease (nLPHD), a 41-year old male developed "B" symptoms and extensive adenopathy. A right axillary lymph node biopsy showed two distinct regions including (1) histiocyte-rich B-cell lymphoma and (2) diffuse small T-cell lymphoma. A clonal rearrangement of the gene for the T-cell receptor beta chain confirmed the presence of a T-cell neoplasm, and this was further confirmed by selective polymerase chain reaction (PCR) on this morphologic zone. PCR on the morphologic B-cell lymphoma confirmed the presence of an immunoglobulin gene rearrangement. These two regions were separated by a less-defined zone containing a mixture of small CD57 positive T lymphocytes, small B lymphocytes, and rare lymphocytic and histiocytic (L&H) cells, highly suggestive of recurrent LPHD. The development of composite B-cell and T-cell lymphoma in this patient raises the speculation that nLPHD may be a neoplasm of lymphoid cells, which can differentiate in both B- and T-cell directions, with the "L&H" cells constituting their B-cell progeny.

Characterization of responses to neurokinin A in the isolated perfused guinea pig heart.

Goals of this study were to identify and characterize effects of neurokinin A (NKA) in isolated guinea pig hearts. Bradycardia, augmentation of ventricular contractions, and reduction of perfusion pressure were prominent responses to bolus injections of NKA (0. 25-25 nmol). NKA was more potent than substance P (SP) in causing bradycardia but did not differ in potency for lowering perfusion pressure. Doses of SP of 25 nmol or less decreased ventricular force, whereas 100 nmol caused a biphasic response. The percent decrease in heart rate produced by 25 nmol NKA was reduced from 58.0 +/- 4.8 to 39.6 +/- 3.5% in the presence of 1 microM atropine (n = 5). The positive inotropic response to 25 nmol of NKA in spontaneously beating hearts was replaced by a negative inotropic response during pacing (22.5 +/- 3.3% increase vs. 11.7 +/- 1.7% decrease, n = 5). Reserpine pretreatment did not affect the positive inotropic response to NKA. Specific binding sites for 125I-labeled NKA were localized to intracardiac ganglia and coronary arteries but not to myocardium. It was concluded that 1) negative chronotropic responses to NKA involve cholinergic and noncholinergic mechanisms, and 2) the positive inotropic response is an indirect action.

Trilineage extramedullary myeloid cell tumor in myelodysplastic syndrome.

A patient with a 17-month history of myelodysplastic syndrome (refractory anemia with excess blasts that evolved into chronic myelomonocytic leukemia), which was treated with transfusions and erythropoietin, developed abdominal and inguinal lymphadenopathy. Biopsies of the abdominal nodes revealed virtual obliteration of the architecture by myeloid blasts admixed with maturing granulocytic, erythroid, and megakaryocytic precursors. The lymph node findings appeared to represent extramedullary dyshematopoiesis undergoing a tissue phase blast transformation. Four months later, the patient developed rising peripheral blast counts consistent with acute leukemia. Although the development of granulocytic sarcoma (also called extramedullary myeloid cell tumor) is well known to occur in patients with myelodysplastic syndromes, to our knowledge this is the first description of an extramedullary myeloid cell tumor associated with trilineage differentiation.

Contribution of hemodynamic variables to the lowering of blood pressure by substance P in the anesthetized rat.

The radioactive microsphere technique was used to determine the contribution of acute changes in systemic hemodynamic variables to the lowering of blood pressure caused by substance P in rats anesthetized with urethane. Infusion of 0.74 nmol/kg/min of substance P caused a decrease of blood pressure, cardiac output, stroke volume and blood flow to most tissues. Total and regional vascular resistances were not affected. Heart rate was increased. These results suggest that the lowering of blood pressure caused by substance P occurs as a result of the decreased stroke volume and cardiac output. The most likely explanation for the decreased stroke volume is a decreased venous return. Several studies have shown that substance P has a direct effect to dilate peripheral arteries. Since substance P dilates arteries, one would expect a decrease of peripheral vascular resistance. The results of this study suggest, however, that counter-regulatory processes, elicited in response to the vasodilatation and direct effects of substance P on sympathetic ganglia to increase the sympathetic nervous system activity, offset the direct effect of substance P on arteries that would otherwise cause a decrease of peripheral vascular resistance.

Pressor and tachycardic responses to intravenous substance P in anesthetized rats.

Intravenous injection of 3-33 nmol/kg of substance P (SP) caused pressor and tachycardic responses in anesthetized rats. The responses were not blocked by a ganglion nicotinic receptor antagonist or by pithing. Pretreatment with reserpine blocked both responses. beta-Adrenoceptor blockade attenuated only the tachycardic response, and alpha-adrenoceptor blockade attenuated only the pressor response. These findings indicated that the effects of SP to increase blood pressure and heart rate are due to sympathetic ganglion stimulation. Studies with adrenalectomized rats showed that stimulation of the adrenals by SP contributes to both responses but makes a greater contribution to the tachycardic response. These observations raise the possibility that the tachykinin innervation of sympathetic ganglia and the adrenal medulla may be involved in the local regulation of blood pressure and heart rate.

Distribution of substance P binding sites in guinea-pig heart and pharmacological effects of substance P.

The localization of substance P (SP) binding sites in guinea-pig heart was studied by in vitro autoradiography, and pharmacological effects of SP were examined with isolated heart preparations. Specific binding of [125I]SP was found in association with cardiac parasympathetic ganglia and some coronary arteries. No specific SP binding sites were associated with coronary veins, atria, ventricles, ascending aorta or pulmonary trunk. Local bolus injections of SP (2.5 and 25 nmol) caused a bradycardia which, in some preparations, was followed by a slight tachycardia. SP produced a prominent coronary vasodilator effect after basal perfusion pressure had been elevated by 1 microM vasopressin. The vasodilator response was probably mediated by the SP binding sites associated with the coronary arteries. Bradycardia might be elicited by binding of SP to the receptors present in the parasympathetic ganglia and subsequent release of acetylcholine. It is suggested that these effects of SP on the isolated heart could be of physiological significance.

Autoradiographic localization of substance P binding sites in guinea-pig airways.

The distribution of substance P (SP) binding sites in guinea-pig airway was examined by in vitro autoradiography with tritium- and iodine-labeled SP. Specific SP binding sites were most abundant in tracheobronchial smooth muscle but were also detected in the mucosa/submucosa. Binding within the mucosa/submucosa was especially high in the region of glands. Binding of iodine-labeled SP to cartilage was negligible. Tritium-labeled SP bound non-specifically to airway cartilage. These observations are consistent with the proposed effects of SP-containing afferent nerves on airway resistance and vascular permeability. The localization of specific SP binding sites suggests that SP may also affect exocrine glands in the respiratory tract.

Distribution of muscarinic receptors and acetylcholinesterase in the rat heart.

Experiments were performed to determine the degree of overlap in the distribution of muscarinic receptors and cholinergic innervation of the rat heart. Localization of muscarinic receptors was determined by autoradiography with [3H]quinuclidinyl benzilate. Adjacent sections were stained for acetylcholinesterase to determine innervation. The distribution of muscarinic receptors and cholinergic innervation overlapped in cardiac parasympathetic ganglia, nodal tissue, His bundle-Purkinje system, vena cava and pulmonary veins. Cholinergic innervation to the right atrium was greater than to the left atrium while muscarinic receptor density was equal in the two atria. Innervation of the ventricles was confined primarily to the base of the right ventricle. A low density of muscarinic receptors was observed throughout the ventricles. Neither cholinergic innervation nor muscarinic receptors were detected in the pulmonary trunk, ascending aorta or cardiac valves. Muscarinic receptors and cholinergic innervation in the nodal regions, ventricular conduction system and myocardium probably mediate negative chronotropic, dromotropic and inotropic effects of vagal nerve stimulation. Muscarinic receptors at sites not containing cholinergic innervation may be associated with noradrenergic nerves of the myocardium.

Effect of vagotomy on cholinergic parameters in nuclei of rat medulla oblongata.

Cholinergic enzymes and muscarinic receptors in nuclei of rat medulla oblongata were examined after unilateral vagotomy to determine their association with efferent vagal neurons. Vagotomy caused an ipsilateral depletion of acetylcholinesterase from the dorsal motor nucleus of the vagus (DNV) and the nucleus ambiguus (NA). Choline acetyltransferase activity was reduced in ipsilateral DNV, nucleus tractus solitarius and rostral NA. Muscarinic receptor localization by autoradiography with [3H]quinuclidinyl benzilate (QNB) revealed marked intranuclear variations in receptor density. Vagotomy had no effect on the QNB binding pattern. Loss of cholinergic enzymes is a consistent response of motor and preganglionic autonomic neurons to axotomy. Depletion of muscarinic receptors is an additional component of axon reaction in brain stem motoneurons. Accordingly, previous studies have shown a decrease in neurotransmitter-related proteins after axotomy of motoneurons. In the present study, cholinergic enzymes were depleted from axotomized vagal neurons but receptors were not. It is concluded that muscarinic receptors in the DNV and NA are not associated with vagal efferent neurons.

Effect of facial nerve transection on acetylcholinesterase, choline acetyltransferase and [3H]quinuclidinyl benzilate binding in rat facial nuclei.

Choline acetyltransferase activity and localization of acetylcholinesterase and [3H]quinuclidinyl benzilate binding sites (muscarinic receptors) in rat facial nuclei were examined 2 weeks after right facial nerve transection or sham control surgery. Choline acetyltransferase activity in the right facial nucleus of nerve-transected rats was only one-third of that in the left nucleus. Histochemical observations revealed loss of acetylcholinesterase from most motoneurons and neuropil of the right facial nucleus after axotomy. Autoradiographic grains, marking muscarinic receptors, were likewise depleted substantially from this region. Facial nuclei of control animals were identical with respect to all of these neurochemical measures and undistinguishable from the left facial nucleus of nerve-transected rats. Cholinergic enzymes are known to be synthesized by motoneurons, but the source of muscarinic receptors in the facial nucleus is not known. Since all three proteins are depleted from the facial nucleus after axotomy of motoneurons, it is concluded that these cells produce cholinergic enzymes and muscarinic receptors. Synthesis of muscarinic receptors by facial motoneurons could indicate these neurons are cholinoceptive. Axotomy should be a useful tool for determining which other neurotransmitter receptors are produced by facial motoneurons and efferent neurons in other cranial nerve nuclei.