Walter Riedel

Regional differentiation of sympathetic activity during hypothalamic heating and cooling in anesthetized rabbits.

SummaryIn anesthetized rabbits immobilized with succinyl choline, the discharges of sympathetic efferents supplying cutaneous and visceral regions were simultaneously recorded. The effects of thermal stimulation of the hypothalamic region were tested on the basis of the integrated discharges. During hypothalamic heating cutaneous sympathetic activity decreased, corresponding to increased ear blood flow, while visceral sympathetic activity increased. During hypothalamic cooling there was, on the average, no significant change of regional sympathetic activity. However, in single experimental periods an increase of cutaneous and a decrease of visceral sympathetic activity was found.The observed responses of regional sympathetic activity were compared with findings about regional cutaneous and intestinal blood flow under the same thermal stimulus and further with corresponding former investigations on regional blood flow and regional sympathetic activity during spinal thermal stimulation. It is suggested by this comparison that regional differentiation of sympathetic activity represents a specific thermoregulatory response of the vasomotor system mediated by the hypothalamic thermoregulatory center.

High-performance liquid chromatographic peak identification of 2,4-dinitrophenylhydrazine derivatives of lipid peroxidation aldehydes by photodiode array detection.

Malonaldehyde (MDA), a product of lipid peroxidation, is a presumptive marker for the development of oxidative stress in tissues and plasmas. In this study we report the photodiode array detection of the 2,4-dinitrophenylhydrazine (DNPH) derivatives of MDA using HPLC. Oxidative stress was produced by injecting (i.p.) bacterial lipopolysaccharide (LPS) into rats at a dose of 100 micrograms/kg, or i.v. into rabbits (1 microgram/kg), or added to freshly drawn human blood (200 ng/ml). Blood was collected at several time points up to 5 h, centrifuged, and equal volumes of 20% TCA were used to precipitate proteins from the plasma. The supernatants were derivatized with DNPH, and the aldehyde-DNPHs were extracted with pentane. After evaporation, aliquots of 10 microliters in acetonitrile were injected onto a Beckman Ultrasphere C18 (3 microns) column, chromatographed with an acetonitrile-water-acetic acid gradient mobile phase and scanned using Waters 996 photodiode array detector. Peak identification and homogeneity was determined by comparing the experimental peaks and UV scans with those of authentic standards. A significant increase in the DNPH derivative of malonaldehyde (MDA-DNPH), but not of the other aldehyde-DNPH derivatives of formaldehyde (FDA), acetaldehyde (ADA), acetone and propionaldehyde (PDA) was seen over the first hour after LPS administration in anesthetized rats, while in conscious rabbits this trend lasted up to 3 h. The retention times as well as the UV scans of the derivatized aldehydes matched the authentic standards. Thus, photodiode array detection has proved valuable in establishing this HPLC method for estimating oxidative stress. This technique could accurately measure pmol amounts of MDA-DNPH indicating the usefulness of photodiode array detection method for estimating small changes in the oxidative stress.

Inhibition of oxygen radical formation by methylene blue, aspirin, or α-lipoic acid, prevents bacterial-lipopolysaccharide-induced fever

Phagocytic cells contain NADPH oxidase that they use for host defense by catalyzing the production of superoxide. Bacterial lipopolysaccharide (LPS) has been found to stimulate NADPH oxidase in mobile and sessile macrophages and microglia. It also evokes fever in homeothermic animals and men, a reaction mediated by central nervous system (CNS) activities. The purpose of the present study was to determine whether reactive oxygen species are involved in LPS-induced fever. In rabbits we found that plasma hydroperoxide levels increased and catalase activity decreased 15 min after LPS injection and that fever started with a similar latency, while plasma levels of tumor necrosis factor-α (TNFα) increased 30 min after the injection. Treating rabbits with methylene blue or aspirin did not affect TNFα secretion but prevented the LPS-induced rise of hydroperoxides and the inactivation of catalase, abolishing fever. Incubation of human blood with nitroblue tetrazolium and LPS increased the number of formazan-positive neutrophils from 10 ± 5 to 52 ± 9%. Adding LPS to blood preincubated with either methylene blue, α-lipoic acid, or aspirin respectively decreased the number of formazan-positive neutrophils to 0.9 ± 0.8, 0.8 ± 0.9, or 2.0 ± 0.9%, disclosing the antioxidant capacity of these drugs. Systemic application of 80 mg/kg α-lipoic acid elicited heat-loss reactions within 15 min and decreased core temperature by 2.2 ± 0.3°C within 2 h. α-Lipoic acid applied 45 min after LPS induced antipyresis within 15 min, and this antipyresis was associated with a decrease of elevated hydroperoxide levels and restoration of catalase activity. Our results show that fever is prevented when the production of reactive oxygen species is blocked and that an elevated body temperature returns to normal when oxygen radical production decreases. Estimation of plasma dihydrolipoic acid (DHLA) levels following injection of 80 mg/kg α-lipoic acid in afebrile and febrile rabbits revealed that this acid is converted into DHLA, which in afebrile rabbits increased the plasma DHLA concentration from 2.22 ± 0.26 μg/ml to peak values of 8.60 ± 2.28 μg/ml DHLA within 30 min and which in febrile rabbits increased it from 0.84 ± 0.22 μg/ml to peak values of 3.90 ± 0.94 μg/ml within 15 min. Methylene blue, aspirin, and α-lipoic acid, which all cross the blood–brain barrier, seem to act not only on peripheral tissues but also on the CNS. Brain structures that have been shown to sense oxidative stress are vicinal thiol groups attached to the NMDA subtype of glutamate receptor. Their reduction by thiol-reducing drugs like dithiothreitol or DHLA has been found to increase glutamate-mediated neuronal excitability, while the opposite effect has been observed after their oxidation. Because we found that systemic application of α-lipoic acid in the afebrile state elicits hypothermia and in the febrile state is antipyretic, we think this type of NMDA receptor is involved in thermoregulation and that oxidation of its thiol groups induces fever. It appears that temperature homeostasis can be maintained only if the redox homeostasis of the brain is guaranteed.

Renal and cutaneous vasomotor and respiratory rate adjustments to peripheral cold and warm stimuli and to bacterial endotoxin in conscious rabbits.

In conscious rabbits peripheral cold stimuli decreased respiratory rate and increased cutaneous vasomotor tone while simultaneously renal sympathetic nervous discharge decreased. Peripheral warm stimuli produced the reverse pattern of autonomic effector activity. Injection of a bacterial endotoxin at warm ambient temperature elicited a biphasic fever response. Within the first 60 min cutaneous vasomotor tone increased, simultaneously renal sympathetic activity decreased. Therefore, ear skin vessels dilated and renal sympathetic activity increased by about 100%. Respiratory rate remained depressed during both fever phases. Renal blood flow was investigated in a second series of experiments and showed a negative stimulation but only in the first phase of fever. The results show that renal innervation is involved in the typical thermoregulatory autonomic activity pattern by which temperature homeostasis is preserved. The vasomotor patterns of cold and heat stress developing during fever are compatible with the concept of a changed set-point of the body thermostat. The depressed respiratory rate and the lack of the renal vascular response to the increased nervous activity during the second fever phase and their reversal to normal after acetylsalicylate (ASA) indicate the participation of prostaglandins modifying peripheral and central neurotransmitter mechanisms.

TUMOR NECROSIS FACTOR-ALPHA IN MACROPHAGES OF HEART, LIVER, KIDNEY, AND IN THE PITUITARY GLAND

Abstract.Tumor necrosis factor-α (TNFα) is an important mediator in bacterial lipopolysaccharide (LPS)-induced fever and shock. New data on TNFα-producing macrophages in heart, pituitary gland, kidneys and liver in correlation with TNFα plasma levels are reported here. In adult rabbits, core temperature and TNFα plasma levels are significantly increased at 3 and 24 h after treatment with LPS. After a delay of 6–12 h, the number of TNFα-containing macrophages, determined by immunohistochemistry, increases more than fivefold in all organs investigated. With the exception of the pituitary gland, the increase in cell number is correlated with the degree of cellular injury, indicating the involvement of TNFα in LPS-induced organ damage that is accompanied by the synthesis of the cytokine. Cortisol levels also increase for at least 24 h after LPS treatment, show peak values 6 h after interleukin-1 treatment, and are unchanged after TNFα treatment, indicating the different effects of these factors on the hypothalamo-hypophyseal-adrenocortical axis. This study provides evidence that macrophageal TNFα of multi-organ origin is involved in LPS-induced tissue injury and supports the concept of a systemic inflammatory response syndrome. We also show for the first time that in the anterior lobe of the pituitary gland TNFα is a normal constituent in cells producing growth hormone but not ACTH. Moreover, most cells of the intermediate lobe are positive for TNFα.

Warm receptors in the dorsal abdominal wall of the rabbit

SummaryWarm stimulation of a circumscribed region of the dorsal abdominal wall of the rabbit evoked discharges in splanchnic afferent single fibers. They showed positive temperature coefficients of their static activities and of their dynamic responses to temperature changes. They were not excited by mechanical stimuli. Cooling from static conditions caused a transient inhibition of impulse frequency. Two populations were found, one having a static and dynamic maximum at 46°C, the other one at about 40°C. Both types were found to be C-fibers, with a conduction velocity of 0.6–1.1 m/sec.

Blunted ACTH and Cortisol Responses to Systemic Injection of Corticotropin-Releasing Hormone (CRH) in Fibromyalgia

Abstract: Thirteen female patients suffering from fibromyalgia (FM) and thirteen female age‐matched controls were intravenously injected with a bolus dose of 100 μg corticotropin‐releasing hormone (CRH), and the evoked secretion pattern of ACTH, cortisol, somatostatin, and growth hormone (GH) was followed up for two hours, together with the plasma levels of CRH. The increases of ACTH and cortisol following CRH were not significantly different between controls and FM patients. The increase of plasma CRH following its injection was significantly higher in FM patients and lasted about 45 min, paralleled by an increase of somatostatin with a similar time course. Basal GH levels were significantly lower in FM patients. GH increased in FM patients 90 min after injection of CRH, coincident with decreasing CRH and somatostatin levels, while GH levels in controls rather decreased with the lowest values occurring 90 min after CRH. The results support the concept that the hormonal secretion pattern frequently observed in FM patients is primarily caused by CRH, possibly as a response to chronic pain and stress. The elevated levels of CRH in the circulation of FM patients suggest elevated levels of CRH‐binding protein, which could explain why the levels of ACTH and cortisol between controls and FM following CRH do not differ.

Hormonal pertubations in fibromyalgia syndrome.

ABSTRACT: The symptomatology characterizing fibromyalgia (FM) comprises three systems: the musculoskeletal system with widespread muscular pain, neuroendocrine disorders, and psychological distress including depression. Though the most prominent symptom of FM is pain in defined points of the musculoskeletal system, the numerous other somatoform and psychological disorders suppose a common primary disturbance which we consider to originate within higher levels of the central nervous system. Recent studies of the entire endocrine profile of FM patients following a simultaneous challenge of the hypophysis with corticotropin‐releasing hormone (CRH), thyrotropin‐releasing hormone, growth hormone‐releasing hormone, and luteinizing hormone‐releasing hormone support the hypothesis that an elevated activity of CRH neurons determines not only many symptoms of FM but may also cause the deviations observed in the other hormonal axes. Hypothalamic CRH neurons thus may play a key role not only in “resetting” the various endocrine loops but possibly also nociceptive and psychological mechanisms as well.

Chapter 6 Role of nitric oxide in temperature regulation

Publisher Summary Nitric oxide (NO) exerts most of its physiological actions by activating a soluble guanylate cyclase as the second messenger, but direct interactions of NO with enzymes and superoxide anions are also considered. NO is assumed to be synthesized in the brain on demand and to leave a neuron or its terminals by diffusion and, therefore, its messenger function must be differently regulated from those of classical transmitters/modulators stored in presynaptic vesicles. The putative role of NO in temperature regulation has only recently attracted interest. In the periphery, the ubiquity of NO generating cells, like the vascular endothelium and the phagocytic elements of the reticulo-endothelial system cells, almost necessarily implies that the widespread actions of NO somehow interfere with thermoregulatory activities. As a ubiquitous systemic mediator, NO has been shown to facilitate thermolytic mechanisms, mainly due to its circulatory action. At the central level, in vivo experiments have suggested a more coordinated function for NO as a thermolytic mediator in thermoregulation and fever. Generally, the distribution of NO producing and NO responsive neurons in central nervous regions of thermoregulatory relevance, like the preoptic and anterior hypothalamus (PO/AH) and the spinal cord, suggests the involvement of NO in temperature signal transmission.

Differentiation of sympathetic activity at the spinal level in Response to central cold stimulation

SummaryThermally induced adjustments of skin blood flow in chronically spinalized dogs may occur without alterations of arterial pressure and heart rate. In lightly anesthetized rabbits with chronic spinal transection at the level of C6/C7, the question was investigated, whether the isolated spinal cord can produce differentiated changes of regional sympathetic activity as a response to central cold stimulation.Selective cooling of the thoraco-lumbar spinal cord in chronically spinalized rabbits induced an increase in vasoconstrictor tone of the ear skin vessels as indicated by a drop of ear skin temperature at constant core and ambient air temperatures. Simultaneously, a decrease of activity in a splanchnic nerve branch was observed. Arterial pressure and heart rate remained unaffected on the average. 24 h after spinal transection, this response seemed to be less pronounced than 3 or more days after the operation. The presented results show that the pattern of regional vasomotor responses induced by central cold stimulation is, on principle, identical in intact and chronically spinalized rabbits. Therefore, the vasomotor pattern described in spinalized animals seems to represent a thermoregulatory response mediated by a vicarious spinal control system.