Intracranial renin alters gustatory neural responses in the nucleus of the solitary tract of rats

Activation of the renin-angiotensin system in the brain is considered important in the arousal and expression of sodium appetite. To clarify the effects of directly activating this hormonal cascade, taste neurons in the nucleus of the solitary tract of rats were tested with a battery of sapid stimuli after intracerebroventricular injection of renin or its vehicle. The rats were chronically prepared but lightly anesthetized during the recording procedure. Eighty-five taste neurons were tested: 46 after renin injections and 39 after vehicle. Neural activity was counted for 5.0-s periods without stimulation (spontaneous) and during stimulation with water and sapid chemicals. The averaged responses to each of the standard stimuli (0.1 M NaCl, 0.3 M sucrose, 0.01 M citric acid, and 0.01 M quinine hydrochloride) did not differ significantly between the two conditions. When the rats were tested with a concentration range of NaCl, however, after renin the average responses to the hypertonic 0.3 and 1.0 M stimuli were reduced to 74 and 70%, respectively, compared with those after vehicle injections. A similar tendency was evident for the subsample of neurons that responded best to NaCl, but the effect was smaller. These data are consistent with, but not as dramatic as, those reported after dietary-induced sodium appetite.     

Leptin in nucleus of the solitary tract alters the cardiovascular responses to aortic baroreceptor activation

Recent data suggests that neurons expressing the long form of the leptin receptor form at least two distinct groups within the caudal nucleus of the solitary tract (NTS): a group within the lateral NTS (Slt) and one within the medial (Sm) and gelantinosa (Sg) NTS. Discrete injections of leptin into Sm and Sg, a region that receives chemoreceptor input, elicit increases in arterial pressure (AP) and renal sympathetic nerve activity (RSNA). However, the effect of microinjections of leptin into Slt, a region that receives baroreceptor input is unknown. Experiments were done in the urethane-chloralose anesthetized, paralyzed and artificially ventilated Wistar or Zucker obese rat to determine leptin's effect in Slt on heart rate (HR), AP and RSNA during electrical stimulation of the aortic depressor nerve (ADN). Depressor sites within Slt were first identified by the microinjection of l-glutamate (Glu; 0.25 M; 10 nl) followed by leptin microinjections. In the Wistar rat leptin microinjection (50 ng; 20 nl) into depressor sites within the lateral Slt elicited increases in HR and RSNA, but no changes in AP. Additionally, leptin injections into Slt prior to Glu injections at the same site or to stimulation of the ADN were found to attenuate the decreases in HR, AP and RSNA to both the Glu injection and ADN stimulation. In Zucker obese rats, leptin injections into NTS depressor sites did not elicit cardiovascular responses, nor altered the cardiovascular responses elicited by stimulation of ADN. Those data suggest that leptin acts at the level of NTS to alter the activity of neurons that mediate the cardiovascular responses to activation of the aortic baroreceptor reflex.     

Calcium deprivation alters gustatory-evoked activity in the rat nucleus of the solitary tract

Calcium-deprived rats develop a compensatory appetite for substances that contain calcium. To investigate the role of gustatory factors in calcium appetite, we recorded the extracellular activity of single neurons in the nucleus of the solitary tract of calcium-deprived and replete rats. The activity evoked by a broad array of taste stimuli was examined in 51 neurons from replete rats and 47 neurons from calcium-deprived rats. There were no differences between the groups in the responses of all neurons combined. However, neurons with sugar-oriented response profiles gave significantly larger responses to 3, 10, and 100 mM CaCl(2) in the calcium-deprived group than did corresponding cells in the replete group. This difference in taste-evoked responding may underlie an increase in the palatability of CaCl(2) and, in turn, contribute to the expression of calcium appetite.     

Acute and chronic exposure to hypoxia alters ventilatory pattern but not minute ventilation of mice overexpressing erythropoietin

Apart from enhancing red blood cell production, erythropoietin (Epo) has been shown to modulate the ventilatory response to reduced oxygen supply. Both functions are crucial for the organism to cope with increased oxygen demand. In the present work, we analyzed the impact of Epo and the resulting excessive erythrocytosis in the neural control of normoxic and hypoxic ventilation. To this end, we used our transgenic mouse line (Tg6) that shows high levels of human Epo in brain and plasma, the latter leading to a hematocrit of approximately 80%. Interestingly, while normoxic and hypoxic ventilation in Tg6 mice was similar to WT mice, Tg6 mice showed an increased respiratory frequency but a decreased tidal volume. Knowing that Epo modulates catecholaminergic activity, the altered catecholaminergic metabolism measured in brain stem suggested that the increased respiratory frequency in Tg6 mice was related to the overexpression of Epo in brain. In the periphery, higher response to hyperoxia (Dejours test), as well as reduced tyrosine hydroxylase activity in carotid bodies, revealed a higher chemosensitivity to oxygen in transgenic mice. Moreover, in line with the decreased activity of the rate-limiting enzyme for dopamine synthesis, the intraperitoneal injection of a highly specific peripheral ventilatory stimulant, domperidone, did not stimulate hypoxic ventilatory response in Tg6 mice. These results suggest that high Epo plasma levels modulate the carotid body's chemotransduction. All together, these findings are relevant for understanding the cross-talk between the ventilatory and erythropoietic systems exposed to hypoxia.     

Hypertension alters GABA receptor-mediated inhibition of neurons in the nucleus of the solitary tract

Previous studies have demonstrated that microinjection of baclofen, a GABA(B) receptor agonist, into the nucleus of the solitary tract (NTS) results in an enhanced pressor response in hypertensive (HT) rats compared with normotensive (NT) rats, suggesting a possible alteration in the responses of neurons in this area to activation of GABA(B) receptors. The following studies were designed to determine whether HT alters the sensitivity of neurons in the NTS to GABA receptor agonists. Sham-operated NT and unilateral nephrectomized, renal-wrap HT Sprague-Dawley rats were anesthetized, and the responses of NTS neurons receiving aortic nerve (AN) afferent inputs to iontophoretic application of GABA, the GABA(A) receptor agonist muscimol, and the GABA(B) agonist baclofen were examined. The AN input was classified as monosynaptic (MSN) if the cell responded to each of two stimuli separated by 5 ms with an action potential. If the cell did not respond, the input was considered polysynaptic (PSN). In MSNs, inhibition of AN-evoked discharge by GABA was not altered in 1 wk of HT but was reduced in 4 wk of HT, whereas in PSNs, sensitivity to GABA was reduced at 1 and 4 wk of HT. In HT rats, inhibition of AN-evoked discharge by baclofen was enhanced in MSNs, but not in PSNs, after 1 and 4 wk of HT, whereas inhibition by muscimol was reduced in MSNs and PSNs at 1 and 4 wk of HT. Changes in sensitivity to muscimol and baclofen within MSNs were the same whether the MSN received a slowly or a rapidly conducted AN afferent input. The results demonstrate that early in HT the sensitivity of NTS neurons to inhibitory amino acids is altered and that these changes are maintained for > or =4 wk. The alterations are dependent on the subtype of GABA receptor being activated and whether the neuron receives a mono- or polysynaptic baroreceptor afferent input.     

Urethane inhibits the GABAergic neurotransmission in the nucleus of the solitary tract of rat brain stem slices

Because urethane is a widely used anesthetic in animal experimentation, in the present study, we evaluated its effects on neurons of the nucleus of the solitary tract (NTS) in brain stem slices from young rats (25-30 days old). Using the whole cell configuration of the patch-clamp technique, spontaneous postsynaptic currents (sPSCs) and evoked excitatory postsynaptic currents (eEPSCs) were recorded. Urethane (20 mM) decreased by approximately 60% the frequency of GABAergic sPSCs (1.0 +/- 0.2 vs. 0.4 +/- 0.1 Hz) but did not change the frequency, amplitude, or half-width of glutamatergic events or TTX-resistant inhibitory sPSCs [miniature inhibitory postsynaptic currents (IPSCs)]. Miniature IPSCs were measured in the presence of urethane plus 1 mM diazepam (1 mM), and no changes were seen in their amplitude. This suggests that the GABA concentration in the NTS synapses is set at saturating level. We also evaluated the effect of urethane on eEPSCs, and no significant change was observed in the amplitude of N-methyl-d-aspartate [NMDA; 44.2 +/- 11.5 vs. 37.6 +/- 10.6 pA (holding potential = 40 mV)] and non-NMDA currents [204.4 +/- 35.5 vs. 196.6 +/- 31.2 pA (holding potential = -70 mV)]. Current-clamp experiments showed that urethane did not alter the action potential characteristics and passive membrane properties. These data suggest that urethane has an inhibitory effect on GABAergic neurons in the NTS but does not change the spontaneous or evoked excitatory responses.     

Responses of neurons of the solitary tract nucleus to noxious colorectal distension in rats

In experiments on anaesthetized rats, the neuronal mechanisms underlying processing of the nociceptive information from the colon within the nucleus of the solitary tract were studied. In addition, the role of nitric oxide in these processes was estimated. Analysis of changes in c-fos expression revealed that nociceptive colorectal distension (CRD) resulted in activation of neurons mainly in the medial, commissural, parvicellular and dorsomedial subnuclei of the solitary tract nucleus. Non-noxious CRD evoked in these subdivisions weak phasic excitatory neuronal responses. Under noxious CRD, neurons with phasic (58%) and tonic (42%) responses were revealed. The phasic neuron responses were significantly enhanced in comparison with non-noxious CRD. Inhibition of the neuronal NO-syntheses resulted in significant decrease of neuron responses to noxious CRD and the number of cells with tonic reactions. Therefore, neurons with tonic responses may be directly related to NO-depended processing ofnociceptive information from colon.     

Hindlimb unloading alters nitric oxide and autonomic control of resting arterial pressure in conscious rats

After periods of microgravity or bed rest, individuals often exhibit reduced Vo(2 max), hypovolemia, cardiac and vascular effects, and autonomic dysfunction. Recently, alterations in expression of vascular and central nervous system NO synthase (NOS) have been observed in hindlimb-unloaded (HU) rats, a model used to simulate physiological effects of microgravity or bed rest. We examined the effects of 14 days of hindlimb unloading on hemodynamic responses to systemic NOS inhibition in conscious control and HU rats. Because differences in NO and autonomic regulation might occur after hindlimb unloading, we also evaluated potential differences in resting autonomic tone and effects of NOS inhibition after autonomic blockade. Administration of nitro-L-arginine methyl ester (L-NAME; 20 mg/kg iv) increased mean arterial pressure (MAP) to similar levels in control and HU rats. However, the change in MAP in response to L-NAME was less in HU rats, that had an elevated baseline MAP. In separate experiments, atropine (1 mg/kg iv) increased heart rate (HR) in control but not HU rats. Subsequent administration of the ganglionic blocker hexamethonium (30 mg/kg iv) decreased MAP and HR to a greater extent in HU rats. Administration of L-NAME after autonomic blockade increased MAP in both groups to a greater extent compared with intact conditions. However, the pressor response to L-NAME was still reduced in HU rats. These data suggest that hindlimb unloading in rats reduces peripheral NO as well as cardiac parasympathetic tone. Along with elevations in sympathetic tone, these effects likely contribute to alterations in vascular control and changes in autonomic reflex function following spaceflight or bed rest.     

Protein kinase C modulation of ventilatory response to hypoxia in nucleus tractus solitarii of conscious rats

This study aimedto determine the role of protein kinase C (PKC) in signal transductionmechanisms underlying ventilatory regulation in the nucleus tractussolitarii (NTS). Microinjection of phorbol 12-myristate 13-acetate intothe commissural NTS of nine chronically instrumented, unrestrained ratselicited significant cardiorespiratory enhancements that lasted for atleast 4 h, whereas administration of vehicle(n = 15) or the inactive phorbol ester 4-phorbol 12,13-didecanoate (n = 7)did not elicit minute ventilation (E)changes. Peak hypoxic Eresponses (10% O₂-balanceN₂) were measured in 19 additional animals after NTS microinjection of bisindolylmaleimide(BIM) I, a selective PKC inhibitor (n = 12), BIM V (inactive analog; n = 7),or vehicle (Con; n = 19). In Con,E increased from 139 ± 9 to 285 ± 26 ml/min in room air and hypoxia, respectively, and similarresponses occurred after BIM V. BIM I did not affect room airE but markedly attenuated hypoxia-induced E increases (128 ± 12 to 167 ± 18 ml/min; P < 0.02 vs. Con and BIM V). When BIM I was microinjected into the cerebellum(n = 4), cortex(n = 4), or spinal cord(n = 4),E responses were similar to Con.Western blots of subcellular fractions of dorsocaudal brain stemlysates revealed translocation of PKC, , , , , and  isoenzymes during acute hypoxia, and enhanced overall PKC activity wasconfirmed in the particulate fraction of dorsocaudal brain stem lysatesharvested after acute hypoxia. These studies suggest that, in the adultrat, PKC activation in the NTS mediates essential components of theacute hypoxic ventilatory response.

     

Chronic sustained and intermittent hypoxia reduce function of ATP-sensitive potassium channels in nucleus of the solitary tract

Activation of neuronal ATP-sensitive potassium (K(ATP)) channels is an important mechanism that protects neurons and conserves neural function during hypoxia. We investigated hypoxia (bath gassed with 95% N(2)-5% CO(2) vs. 95% O(2)-5% CO(2) in control)-induced changes in K(ATP) current in second-order neurons of peripheral chemoreceptors in the nucleus of the solitary tract (NTS). Hypoxia-induced K(ATP) currents were compared between normoxic (Norm) rats and rats exposed to 1 wk of either chronic sustained hypoxia (CSH) or chronic intermittent hypoxia (CIH). Whole cell recordings of NTS second-order neurons identified after 4-(4-(dihexadecylamino)styryl)-N-methylpyridinium iodide (DiA) labeling of the carotid bodies were obtained in a brain stem slice. In Norm cells (n = 9), hypoxia (3 min) induced an outward current of 12.7 +/- 1.1 pA with a reversal potential of -73 +/- 2 mV. This current was completely blocked by the K(ATP) channel blocker tolbutamide (100 muM). Bath application of the K(ATP) channel opener diazoxide (200 muM, 3 min) evoked an outward current of 21.8 +/- 5.8 pA (n = 6). Hypoxia elicited a significantly smaller outward current in both CSH (5.9 +/- 1.4 pA, n = 11; P < 0.01) and CIH (6.8 +/- 1.7 pA, n = 6; P < 0.05) neurons. Diazoxide elicited a significantly smaller outward current in CSH (3.9 +/- 1.0 pA, n = 5; P < 0.05) and CIH (2.9 +/- 0.9 pA, n = 3; P < 0.05) neurons. Western blot analysis showed reduced levels of K(ATP) potassium channel subunits Kir6.1 and Kir6.2 in the NTS from CSH and CIH rats. These results suggest that hypoxia activates K(ATP) channels in NTS neurons receiving monosynaptic chemoreceptor afferent inputs. Chronic exposure to either sustained or intermittent hypoxia reduces K(ATP) channel function in NTS neurons. This may represent a neuronal adaptation that preserves neuronal excitability in crucial relay neurons in peripheral chemoreflex pathways.     

Subthreshold aortic nerve inputs to neurons in nucleus of the solitary tract

Subthreshold aortic nerve (AN) inputs to neurons receiving a monosynaptic AN-evoked input (MSNs: respond to each of two AN stimuli separated by 5 ms) and neurons receiving a polysynaptic AN input (PSNs) in the nucleus of the solitary tract (NTS) were identified in anesthetized rats. In extracellular recordings from 24 MSNs and 49 PSNs, 12% of MSNs and 29% of PSNs only responded to AN stimulation during the application of excitatory amino acids. In intracellular recordings from 24 MSNs and 22 PSNs, 12% of MSNs and 14% of PSNs responded to AN stimulation with excitatory postsynaptic potentials that did not evoke action potential discharge. Reductions in arterial pressure produced minimal changes in the spontaneous discharge of suprathreshold AN-evoked neurons, suggesting that these neurons receive excitatory inputs from nonbaroreceptor sources. The results suggest that some baroreflex-related NTS neurons exist in a "reserve state and can be changed to an active state or vice versa. This will change the number of neurons involved in baroreflex circuits and provides a novel mechanism for regulating baroreflex function independently of alterations in peripheral afferent input.     

Respiratory responses to microinjections of leptin into the solitary tract nucleus

Regulatory polypeptide leptin, apart from its well-known hypothalamic effects, stimulates ventilation. The present study on anaesthetised rats was undertaken to elucidate the respiratory effects of 10(-10)-10(-4) M leptin microinjected into the solitary tract nucleus, containing a high concentration of leptin receptors. Injections of 10(-8)-10(-4) M leptin induced dose-dependent increase in ventilation, tidal volume and electric activity of inspiratory muscles; 10(-6) M leptin additionally induced a short-term increase in respiratory frequency and a shortening of both inspiratory and expiratory duration. The respiratory responses to leptin is also characterised by appearance of sighs: deep and prolonged inspirations associated with an augmented burst in the activity of the inspiratory muscles and prolonged post-sigh inter-burst interval. The results taken together with evidence of high concentration of specific leptin ObRb-receptor in the solitary tract nucleus suggest involvement of endogenous leptin in the control of breathing via dorsal structures of the respiratory center.     

Effect of intermittent chronic exposure to hypoxia on feeding behaviour of rats

Healthy albino male rats were exposed to a simulated high altitude (HA) equivalent to 25000 ft (7620 m) for 6 h daily, continuously for 21 days to study the feeding behaviour. The 24-h food and water intake and body weight once in 3 days were recorded. Blood samples were drawn once a week from the retro-orbital venous plexus for blood sugar analysis. All the parameters were recorded before, during and after exposure to simulated HA. The results show a decrease in 24-h food and water intake and decreased gain in body weight during hypoxic exposure, which showed a tendency to come back to control during the post-exposure period. The blood sugar reflected a state of mild hyperglycaemia during exposure to HA.     

Dietary NaCl influences the organization of chorda tympani neurons projecting to the nucleus of the solitary tract in rats

Prior research has shown that maintained exposure to either a low or high NaCl diet from conception to adulthood is associated with changes in NaCl solution intake and neural responses of the chorda tympani (CT) nerve. The present study examined the influence of maintained exposure to a low or high NaCl diet on the central organization of CT neurons projecting to the nucleus of the solitary tract (NST). Three groups of rats were reared and maintained on regular chow containing either basal 0.1%, intermediate 1.0% or high 6% NaCl from conception to adulthood. The fluorescent marker Dil was applied to the CT for characterization of afferent terminations and efferent cell body labeling in the brainstem. The total NST area occupied by CT afferent fibers was the same for all three dietary groups. However, the pattern of CT innervation differed such that there was an enlarged dorsal terminal field in the high group. There were no group differences in body and brain weight, or in efferent labeled neurons. Thus, Dil has been demonstrated to be an effective transport marker of the gustatory system and the parameters of dietary NaCl exposure that influence the pattern of the CT fibers projecting to the NST have been further clarified.     

Role of the solitary tract nucleus and caudal ventrolateral medulla in temperature responses in endotoxemic rats

In experiments on conscious rats it was found that preliminary microinjection of 100 nl 100 microM glutamic acid to the rostral commissural part of the solitary tract nucleus or to the caudal ventrolateral medulla increased a rise in colonic temperature induced by systemically applied endotoxin (3 microg/kg Escherichia coli lipopolysaccharide, i.p.) as compared to animals with intrabulbar injection of vehicle (control group). Preliminary microinjection of glutamate to the caudal commissural part of the solitary tract nucleus levelled the endotoxin-induced temperature response. After glutamate treatment of the caudal ventrolateral medulla there was a significant decrease in the noradrenaline content and decrease in the adrenaline level in the caudal (not significant) and rostral ventrolateral medulla (significant), as well as a small rise in noradrenergic activity at the solitary tract nucleus as compared to control animals. The post-mortem measurement of the optical density of brainstem tissues revealed its significant attenuation at the solitary tract nucleus and caudal ventrolateral medulla after glutamate as compared with these structures after vehicle. The involvement of monoaminergic systems of both structures under study in the initiation and control of temperature responses during endotoxemia is suggested.     

Respiratory responses to microinjections of gastrin-releasing peptide into the solitary tract nucleus

In acute experiments on urethane-anesthetized rats, the respiratory effects ofmicroinjections of 10(-5), 10(-8) and 10(-10) M gastrin-releasing peptide (GRP) into the solitary tract nucleus were investigated. It was found that microinjections of the neuropeptide induced an increase in tidal volume, amplitude of diaphragm and external intercostal muscles firing activity and in expiratory duration. The most obvious respiratory responses observed when 10(-8) M GRP was used, while 10(-10) M GRP appeared to be sub-threshold and didn't alter the breathing pattern and activity of inspiratory muscles. In some experiments, where the blood pressure and the heart rate was monitored alone with breathing pattern, these parameters did not change after GRP microinjections into the solitary tract nucleus. The obtained data together with particularities of the distribution of GRP receptors in the brainstem suggest the possibility of GRP involvement into the respiratory control mechanisms at the level of solitary tract nucleus.