Exposure to alcohol in utero blunts splenic sympathetic neural response to endotoxin and interleukin-1β in the rat

Systemic administration of endotoxin (LPS) or interleukin-1β (IL-1) to prepubertal rats induced a marked increase in splenic but not cardiac norepinephrine (NE) turnover, an index of sympathetic neural activity. In contrast, the splenic neural response was blunted in their fetal alcohol-exposed (FAE) cohorts. Because the sympathetic outflow to lymphoid organs is considered an important immune modulator, the anomalous neural response to immune signals may partly account for the impaired cellular immunity and, thus, for the increased susceptibility to infections associated with FAE.     

Cytokine-induced change in hypothalamic norepinephrine turnover: involvement of corticotropin-releasing hormone and prostaglandins

Changes in norepinephrine (NE) turnover in restricted brain regions were examined in rats after administration of the major mediators of the acute phase response, interleukin-1β (IL-1), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF). An increase in NE turnover was observed after intraperitoneal injection of IL-1 (1 μg/rat) in the whole hypothalamus and several specific hypothalamic nuclei, but not in the medulla oblongata and cerebral cortex. The stimulatory effect of IL-1 was mimicked by an intracerebroventricular injection of much lower doses of IL-1 (10–100 ng/rat). This IL-1-induced increase in hypothalamic NE turnover was blocked by the pretreatment with either indomethacin (cyclooxygenase inhibitor) or anti-corticotropin releasing hormone (CRH) antibody but not by naloxone. Intracerebroventricular injection of CRH increased NE turnover not only in the hypothalamus but also in the medulla oblongata and cerebral cortex. However, prostaglandin (PG) E₂ and PGF_2α did not show such effect. It was therefore suggested that IL-1 activates noradrenergic neurons projecting to the hypothalamus by its direct action to the brain, and that CRH and eicosanoid-cyclooxygenase product(s) within the brain are involved in this process. In contrast, neither IL-6 nor TNF influenced brain NE turnover regardless of whether they were given intraperitoneally or intracerebroventricularly. Thus, although IL-6 and TNF, as well as IL-1, show common central effects such as fever and pituitary-adrenal activation, these effects may be independent of the activation of NE metabolism in the hypothalamus.     

Evidence for a different sensitivity to various central effects of interleukin-1 β in mice

Interleukin 1 (IL-1) induces a series of metabolic and endocrine effects. Activation of the hypothalamus-pituitary-adrenal axis, inhibition of food and water intake, elevation of serum interleukin-6 (IL-6) concentration and hypoglycemia are some of the effects induced by IL-1. The purpose of this study was to compare the sensitivity of these effects following central and peripheral administration of IL-1β. Different doses of IL-1β (0.1–1000 ng/mouse) were centrally (ICV) or peripherally (IP) injected to male mice two hours prior to sacrifice. The ICV administration was more efficacious than the IP injection in elevating serum corticosterone and IL-6 concentrations, whereas no difference was evident in the IL-1β-induced hypoglycemia. Central IL-1β administration was also more potent than IP injection in inhibiting overnight food and water intake. A dose-dependent effect was evident in all these cases. In summary, our data compare effects elicited by central or peripheral administration of different doses of IL-1β. This comparison suggests that the IL-1β stimulation of serum corticosterone and IL-6 and inhibition of food and water intake are events more centrally mediated than the IL-1β-induced hypoglycemia.     

Via beta-adrenoceptors, stimulation of extrasplenic sympathetic nerve fibers inhibits lipopolysaccharide-induced TNF secretion in perfused rat spleen

Using a spleen slice microsuperfusion technique in mice, we have previously characterized the role of norepinephrine (NE) and other neurotransmitters for sympathetic modulation of IL-6 and TNF secretion of splenic macrophages. Since experiments in spleen slices do not reflect the situation of an entire perfused organ, we investigated sympathetic modulation of lipopolysaccharide (LPS)-induced secretion of IL-6 and TNF in perfusion experiments of rat spleen. In an organ bath, perfusion was performed in explanted whole spleens, and catecholamines and cytokines were measured by HPLC and ELISA, respectively. Release of NE depended on stimulation frequency (maximum at 10 Hz). Apart from NE, perfusates also contained significant amounts of dopamine and epinephrine. Furthermore, perfusate epinephrine levels correlated positively with perfusate NE levels (RRank=0.750, p<0.001) but not with plasma epinephrine concentrations. This indicates that epinephrine is a conversion product of sympathetically released NE. Early electrical stimulation of extrasplenic splenic nerves, 20 min after administration of LPS, significantly inhibited TNF secretion. This electrically induced effect was abrogated by simultaneous administration of propranolol (10(-6) M) but it was not influenced by administration of either an alpha1- or alpha2-adrenergic antagonist. Late electrical stimulation of splenic nerves, 2.5 h after administration of LPS, did not change TNF secretion. Interestingly, no influence of early or late sympathetic nerve fiber stimulation on IL-6 secretion was observed. In conclusion, this is the first perfusion study of the entire spleen that demonstrates that early electrical stimulation of sympathetic splenic nerve fibers directly inhibits LPS-induced TNF secretion. This study corroborates the idea that splenic sympathetic nerves are able to inhibit important activators of the innate immune system when stimulation happens very early or even prior to their induction by LPS.     

Intracerebral interleukin-1β impairs response to tumor invasion: involvement of adrenal catecholamines

Interleukin-1β (IL-1β) is released within the brain following stress, trauma, infection, and in specific brain disorders. This centrally acting IL-1β has recently been shown to impair peripheral immunity. Central administration of IL-1β suppresses natural killer (NK) cell activity impairs lung clearance of tumor cells and enhances tumor colonization. Using an in vivo model of tumor colonization (lung clearance of NK-sensitive MADB106 adenocarcinoma cells), this study examined the role of the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system (SNS) in mediating these effects. We demonstrate that adrenalectomy significantly attenuated the impaired lung clearance of MADB106 tumor cells induced by intracerebroventricular (i.c.v.) administration of IL-1β (20 ng). Supplementing adrenalectomized animals with corticosterone did not reinstate the effect. The effect of IL-1β on lung clearance was blocked by pretreatment with the β-adrenergic antagonist, nadolol (0.5 mg/kg), but not by the α-antagonist phentolamine (5 mg/kg). Peripheral noradrenergic pathways are not implicated given that systemic administration of the noradrenergic neurotoxin, 6-hydroxydopamine, did not block the effect of IL-1β. Taken together, these findings indicate that IL-1β impairs lung clearance of MADB106 tumor cells via the actions of adrenal catecholamines, most likely epinephrine, acting at β-adrenergic receptors in the periphery.     

Alterations in T lymphocyte activity following chemical sympathectomy in young and old Fischer 344 rats

In aged Fischer 344 (F344) rats, sympathetic noradrenergic (NA) innervation of the spleen is markedly diminished compared with young rats. To determine if diminished NA innervation can still provide functional signals to splenic T cells, young (3 months old) and old (17 months old) F344 rats were treated with the NA-selective neurotoxin, 6-hydroxydopamine (6-OHDA) to destroy peripheral NA nerve fibers. In 3-month-old rats, no alterations in spleen cell Con A-induced T cell proliferation, IL-2 or IFN-gamma production were observed up to 15 days after sympathectomy, when splenic NE was maximally depleted. By 21 days post-sympathectomy, when NE levels had partially recovered, Con A-induced proliferation and IFN-gamma production, but not IL-2 production, were reduced in sympathectomized animals. After day 21 post-sympathectomy, no alterations in T cell functions were observed in sympathectomized animals. In 17-month-old rats, spleen cell Con A-induced proliferation and IL-2 production were reduced 5 days after sympathectomy in the absence of changes in CD5+ T cells or IFN-gamma production. Desipramine pretreatment, to block 6-OHDA uptake and prevent sympathectomy, completely blocked the 6-OHDA-induced effects, demonstrating that the destruction of NA nerve fibers is required. After day 5 post-sympathectomy, no sympathectomy-induced alterations in Con A-induced T cell functions were observed in old animals. These differences between young and old rats demonstrate that old animals are more susceptible to loss of sympathetic NA innervation, perhaps because compensatory mechanisms are limited. The sympathectomy-induced reduction in T cell proliferation indicates that splenic NA innervation in old animals, though diminished, can exert a positive regulatory influence on T lymphocyte function. Further study of sympathetic neural-immune interactions in the aged rat may provide a means to improve T cell responsiveness in aging.     

Memory impairment induced by IL-1β is reversed by α-MSH through central melanocortin-4 receptors

Interleukin-1beta (IL-1β) significantly influences memory consolidation. Treatments that raise the level of IL-1β in the brain, given after training, impair contextual fear conditioning. The melanocortin α-MSH exerts potent anti-inflammatory actions by physiologically antagonizing the effect of pro-inflammatory cytokines. Five subtypes of melanocortin receptors (MC1R–MC5R) have been identified, with MC3R and MC4R predominating in the central nervous system. The present experiments show that injection of IL-1β (5 ng/0.25 μl) in dorsal hippocampus up to 15 min after training decreased freezing during the contextual fear test. The treatment with IL-1β (5 ng/0.25 μl) 12 h after conditioning cause amnesia when animals were tested 7 days post training. Thus, our results also demonstrated that IL-1β can influence persistence of long-term memory. We determined that animals previously injected with IL-1β can acquire a new contextual fear memory, demonstrating that the hippocampus was not damaged. Treatment with α-MSH (0.05 μg/0.25 μl) blocked the effect of IL-1β on contextual fear memory. Administration of the MC4 receptor antagonist HS014 (0.5 μg/0.25 μl) reversed the effect of α-MSH. However, treatment with γ-MSH (0.5 μg/0.25 μl), an MC3 agonist, did not affect IL-1β-induced impairment of memory consolidation. These results suggest that α-MSH, through central MC4R can inhibit the effect of IL-1β on memory consolidation.     

In vitro superfusion method for the investigation of nerve-immune cell interaction in murine spleen

Immune defence mechanisms can be modulated by brain function. To study such interactions, an in vitro method was developed to examine the release of cytokines and norepinephrine (NE) after electrical stimulation. Slices of mouse spleen were placed in chambers with a volume of 80 μl and superfused with culture medium. To characterize electrically evoked NE release and cell viability a suitable stimulation protocol was developed using of [³H]NE. As parameter for immune function, modulation of interleukin-6 (IL-6) release by the spleen cells brought about by electrical stimulation was investigated. Splenic [³H]NE overflow was calcium-dependent, tetrodotoxin-sensitive and elicited by 54 mM potassium. Electrically evoked [³H]NE release at 22 h was about 80% of the release at 5.3 h. Electrical stimulation substantially reduced IL-6 secretion at 6 h (control: 143.4 ± 14.3 vs. electrical: 71.3 ± 7.9 pg/ml/10⁶ leukocytes, P = 0.0001). This effect was inhibited in a concentration-dependent manner by the β-adrenergic antagonist propranolol (P = 0.0298, EC₅₀approx. 10⁻⁷M). In conclusion, this new technique allows long-term investigation of cell function in slices of murine spleen. In addition, these are the first in vitro data indicating the presence of a functional neuroimmunological link in murine lymphoid tissue.     

Neuropeptide specificity of prostaglandin E2-induced activation of splenic and renal sympathetic nerves in the rat

Sympathetic activation occurs rapidly following intracerebroventricular (icv) injection of prostaglandin E2(PGE2). This study examined whether neuropeptides mediate PGE2-induced sympathetic nerve activation in urethane/chloralose-anesthetized Sprague-Dawley rats. Animals were pretreated (20.0 microg, icv) with the following receptor antagonists; CRF ([D-Phe12,Nle21,38,Calpha-MeLeu37]CRF12-41), AVP-V1 (Des-Gly-[Phaa1, D-Tyr(Et)2,Lys6,Arg8]-vasopressin), or OT (OT+V1, [d(CH2)5,Tyr(Me)2,Orn8]-vasotocin) followed 20 min later by PGE2 (2.0 microg, icv). Pretreatment with the CRF antagonist attenuated the increase in renal nerve activity induced by PGE2 when measured 10 and 30 min post-injection. PGE2-induced renal nerve activity was also inhibited at both time points by the AVP antagonist and, to a similar extent, the OT antagonist. The AVP antagonist did not effect splenic nerve responses to PGE2 whereas the CRF antagonist produced an incomplete and transient reduction in PGE2-induced activation of the splenic nerve. However, the OT antagonist completely blocked the activation of the splenic nerve after central injection of PGE2. ICV injections of AVP and OT produced immediate changes in splenic and renal nerve activity whereas CRF failed to alter the activity of either nerve in anesthetized or conscious animals. Thus, PGE2 acts through neuropeptide-specific pathways to initiate sympathetic outflow and OT is a specific component of the sympathetic pathway innervating the spleen.     

Effects of a chronic inflammatory stress on levels of pro-opiomelanocortin-derived peptides in the rat spleen and thymus

Adjuvant-induced arthritis (AA) in specific strains of rats is an immunologically mediated inflammatory disease which is also characterised by activation of the endocrine system. To further investigate the effects of AA on processing of the pro-opiomelanocortin (POMC) precursor in rat immune tissues, we utilised radioimmunoassays for adrenocorticotropin (ACTH), β-endorphin and α-melanocyte-stimulating hormone (α-MSH) to measure these peptides in the spleen and thymus. 14 days following adjuvant injection, spleen levels of ACTH were elevated in the AA group (4.47 ± 1.04 ng/g tissue, n = 9) compared to controls 2.42±0.4 ng/g) and exacerbation of the disease by removal of circulating glucocorticoids through bilateral adrenalectomy (ADX) resulted in further elevation of spleen ACTH (5.11 ± 1.22 ng/g). β-Endorphin levels in both the AA (10.60 ± 1.61 ng/g) and AA/ADX (13.37 ± 2.36 ng/g) groups were higher than controls 5.57 ± 0.65 ng/g). Conversely, α-MSH spleen levels were decreased in the AA (2.89 ± 0.22 ng/g) and AA/ADX (2.22 ± 0.33 ng/g) groups compared to controls (4.62 ± 0.45 ng/g) and were also decreased following adrenalectomy. In the thymus, ACTH levels were elevated in the AA group (8.95 ± 1.41 ng/g) compared to controls (5.79 ± 0.63 ng/g), and the same pattern wasevident for thymic α-MSH (0.64 ± 0.08 ng/g in AA animals compared to control levels of 0.35 ± 0.03 ng/g). Following G50 gel filtration, ACTH and β-endorphin immunoreactivities (ir) were present in both spleen and thymus as two peaks, one which eluted near the void volume and one which eluted in a lower molecular mass position than the standards. However, ir-ACTH and ir-β-endorphin were present in multiple peaks in AA spleen extracts, indicating extensive processing. These data are the first evidence that ir-ACTH and ir-β-endorphin tissue content and POMC processing in the spleen and thymus can be influenced by a chronic stress.     

Blockade of angiotensin II AT1 receptors inhibits pressor action of centrally administered interleukin-1β in Sprague Dawley rats

The aim of the present study was to evaluate the influence of intraventricular administration of recombinant rat interleukin-1β (IL-1β) on regulation of resting blood pressure and heart rate and to test the hypothesis that the brain angiotensinergic system is involved in regulation of hemodynamic parameters by centrally applied IL-1β. The experiments were performed on Sprague Dawley rats, assigned to three series of experiments. In series 1 (control, n = 6), mean arterial pressure (MAP) and heart rate (HR) were recorded for 15 min under baseline conditions. This was followed by infusion of saline (0.9% sterile NaCl 5 μL/h) into the left cerebral ventricle (LCV). Measurements were continued during the next 60 min. In series 2 (n = 6) and 3 (n = 6) the experimental design was similar, except that in series 2 the animals were LCV infused with saline containing IL-1β (100 ng/h) and in series 3 with saline containing IL-1β (100 ng/h) and angiotensin type 1 (AT1) receptors antagonist (Losartan, 10 μg/h). LCV infusion of saline alone did not influence MAP and HR while administration of IL-1β elicited significant increase in MAP, but not in HR. The pressor effect was absent during combined infusion of IL-1β and Losartan. Results of the study provide evidence that centrally administered IL-1β exerts pressor effect, and reveal that this effect is mediated by stimulation of the brain angiotensin system and requires activation of AT1 receptors.     

HIV-1 Envelope Glycoprotein 120 Regulates Brain IL-1β System and TNF-α mRNAs In Vivo

Human immunodeficiency virus type I (HIV-1)-derived envelope glycoprotein 120 (gp120) is proposed to play an important role in HIV-1 neuropathology. Gp120 may act through mediators including proinflammatory cytokines. Here, we investigated the regulation of the IL-1β system [IL-1β, IL-1 receptor type I (IL-1RI), IL-1 receptor antagonist (IL-1Ra)], TNF-α and TGF-α mRNAs in the rat central nervous system (CNS) in response to the constant intracerebroventricular (ICV) microinfusion of HIV-1 gp120 for 72 h and 144 h. The results show that gp120: (1) increased IL-1β and IL-1Ra mRNAs levels in the same samples from the cerebellum, hypothalamus and midbrain, with the largest increase in the hypothalamus; (2) induced profiles of IL-1β mRNA and IL-1Ra mRNA that were highly intercorrelated; (3) increased the hypothalamic TNF-α mRNA levels; and (4) did not affect the IL-1RI mRNA and TGF-α mRNA levels in any brain region. A dysregulation in the IL-1β/IL-1Ra CNS balance and a mutual induction and synergistic activity of IL-1β and TNF-α could result in a deleterious amplification cycle of cellular activation and cytotoxicity with implications to HIV-1-associated encephalitis, encephalopathy, and neurological manifestations.     

β-Endorphin-induced increases in plasma epinephrine, norepinephrine and dopamine in rats: inhibition of adrenomedullary response by intracerebral somatostatin

Synthetic human β-endorphin, 7.25 nmol intracisternally, in unanesthetized, freely moving, chronically cannulated, adult male rats increased plasma concentrations of all 3 catecholamines: epinephrine, norepinephrine and dopamine, for the 2 h period studied. Blockade of these endorphin effects by the prior systemic administration of naloxone supports mediation of the effects at opioid receptors. Acute systemic administration of guanethidine, which decreases norepinephrine release induced by sympathetic nerve stimulation, blunted the plasma norepinephrine response to intracerebral β-endorphin. Thus, it seems likely that in addition to secretion by adrenal medulla a considerable portion of the β-endorphin-induced increase in norepinephrine is derived from sympathetic nerve endings. Simultaneous intracisternal administration of another neuropeptide, somatostatin, together with β-endorphin markedly inhibited the plasma epinephrine response to β-endorphin, while decreasing the dopamine and norepinephrine responses to a much lesser degree. The data suggest that β-endorphin stimulates central sympathetic outflow to both adrenal medulla and sympathetic nerve endings, and further that somatostatin inhibits the effect of endorphin to stimulate outflow to adrenal medulla but does not affect outflow to sympathetic nerve endings.     

Interleukin-6 production by astrocytes: Induction by the neurotransmitter norepinephrine

Astrocytes contribute to the immunocompetence of the central nervous system (CNS) via their expression of class II major histocompatibility complex (MHC) antigens and the production of inflammatory cytokines such as interleukin-1 beta (IL-1β), tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6). Of these cytokines, IL-6 is of particular interest because one of its many immune and inflammatory actions is the promotion of immunoglobulin synthesis, and it is thought that IL-6 expression within the brain exacerbates autoimmune diseases of the CNS, which are marked by local immunoglobulin production. Several stimuli induce astrocyte IL-6 expression, including such inducible endogenous factors as IL-1β and TNF-α. We have investigated the possibility that a constitutively present endogenous factor, the neurotransmitter norepinephrine (NE), can induce astrocyte IL-6 production. We report that NE induces both IL-6 mRNA and protein in primary neonatal rat astrocytes, with optimal induction at 10 μM. IL-6 protein induction by NE is comparable to that seen with IL-1β or TNF-α, and NE synergizes with these cytokines for a ten-fold enhanced effect. In contrast to astrocytes, microglia are relatively unresponsive to NE, IL-1β and TNF-α for IL-6 production. Experiments with the β-adrenergic receptor agonist isoproterenol, and α and β-adrenergic receptor antagonists (propranolol, phentolamine, atenolol, and yohimbine) indicate that β₂ and α₁-adrenergic receptors are involved in NE induction of astrocyte IL-6 expression. These results help to further the understanding of neuron-glial interactions, and the role of astrocytes and adrenergic activity in immune responses within the CNS.     

Re-exposure to endotoxin induces differential cytokine gene expression in the rat hypothalamus and spleen

This study was designed to investigate whether the pattern of hypothalamic and splenic cytokine expression induced by peripheral administration of a bacterial lipopolysaccharide (LPS) is affected by prior exposure to LPS derived from another bacterial strain. Injection of LPS from Salmonella enteritidis (LPS₂) alone resulted in increased hypothalamic gene expression of IL-1β, IL-6, TNFα, IL-1ra and IL-10. However, pre-exposure to LPS derived from Escherichia coli (LPS₁) 3 weeks before, significantly attenuated hypothalamic IL-1ra, IL-6 and IL-10 expression. IL-1β expression also tended to be lower. This pattern contrasted with the robust cytokine expression in the spleen of LPS₂-treated rats previously exposed to LPS₁, since pre-treatment with endotoxin resulted in a significantly greater response of IL-1β and IL-1ra to LPS₂. Expression of TNFα and IL-10 also tended to be higher. Pre-treatment with LPS₁ did not significantly affect the marked increase in corticosterone and adrenaline blood levels induced by LPS₂. Thus, while endotoxin pre-exposure seemed not to induce a “tolerant” state in the periphery as judged by the immune and endocrine parameters evaluated upon re-stimulation, expression of four of the six cytokines measured was decreased in the hypothalamus. This is the first demonstration that endotoxin priming can differentially affect cytokine expression in the central nervous system and peripheral tissues when a host is confronted with a second, acute, pro-inflammatory stimulus. These results may provide new evidence for the involvement of cytokine pathways in the central nervous system in modulating peripheral inflammation and mediating cognitive and behavioural alterations during inflammatory diseases.     

α2-Adrenoceptor-mediated restraint of norepinephrine synthesis, release, and turnover during immobilization in rats

Stress-related release of norepinephrine (NE) in the brain and periphery probably underlies several neuroendocrine and neurocirculatory responses. NE might influence its own synthesis, release, and turnover, by negative feedback regulation via α₂-adrenoceptors. We examined central and peripheral noradrenergic function by measuring concentrations of NE, dihydroxyphenylglycol (DHPG), and dihydroxyphenylacetic acid (DOPAC) in hypothalamic paraventricular nucleus (PVN) microdialysate and arterial plasma simultaneously during immobilization (IMMO) in conscious rats. The α₂-adrenoceptor antagonist yohimbine (YOH) was injected i.p. or perfused locally into the PVN via the microdialysis probe. The i.p. YOH increased plasma NE, epinephrine (EPI), DHPG, dihydroxyphenylalanine, and DOPAC levels by 4.3, 7.3, 2.5, 0.6 and 1.8-fold and PVN microdialysate NE, DHPG, and DOPAC by 1.2, 0.6 and 0.5-fold. The i.p. YOH also enhanced effects of IMMO on plasma and microdialysate NE, DHPG, and DOPAC. YOH delivered via the PVN microdialysis probe did not affect microdialysate or plasma levels of the analytes at baseline and only slightly augmented microdialysate NE responses to IMMO. The results indicate that α₂-adrenoceptors tonically restrain NE synthesis, release, and turnover in sympathetic nerves and limit IMMO-induced peripheral noradrenergic activation. In the PVN, α₂-adrenoceptors do not appear to contribute to these processes tonically and exert relatively little restraint on IMMO-induced local noradrenergic activation.     

Focal traumatic brain injury causes widespread reductions in rat brain norepinephrine turnover from 6 to 24 h

The effect of right sensorimotor traumatic brain injury (TBI) in male Sprague-Dawley rats on brain norepinephrine (NE) turnover was assessed by measuring the decline of endogenous NE levels following tyrosine hydroxylase inhibition produced with α-methyl-p-tyrosine. Right sensorimotor cortex contusions were produced by a pneumatically driven piston which depressed the dural surface by 2 mm at 3.2 m/s. TBI rats were compared to uninjured, anesthetized controls at 6 h and 24 h after surgery. While NE turnover was not affected at the lesion site at 6 h after TBI, it was either abolished or decreased by 33–75% bilaterally in the hypothalamus and in the cerebral cortex surrounding and rostral to the lesion site. In the cortex caudal to the lesion site, NE turnover was completely abolished. NE turnover in cerebral cortex opposite the lesion site and in the contralateral cerebellum was decreased by 51 and 43%, respectively, at 6 h. At 24 h, NE turnover was either abolished or decreased bilaterally by 45–92% in all cortical areas, in the hypothalamus, cerebellum, locus coeruleus and medulla. Thus, right sensorimotor cortex contusion causes a marked, early and widespread depression of brain NE turnover. Since amphetamine increases NE turnover, this may explain the dramatic improvement in behavioral deficits which occurs following amphatamine administration at 24 h after such lesions.     

Limited responsiveness of LHRH neurons norepinephrine may account for failure of locus coeruleus or medullary A1 electrical stimulation to increase plasma LH in estrogen-treated ovariectomized rats

We recently reported that electrical stimulation (ES) of the locus coeruleus (LC) or the medullary A1 noradrenergic cell groups markedly increased LH secretion. However, these amplifying effects occurred only in rats in which preliminary electrochemical stimulation (ECS) of the medial preoptic nucleus (MPN) was performed. In contrast, ES of either LC or A1 alone did not alter basal LH secretion. Possible explanations for this dichotomy in LH response include: (1) LHRH neurons in unanesthetized, estrogen-treated, ovariectomized (OVX) rats are relatively unresponsive to NE, (2) the chloral hydrate anesthesia used in our brain stimulation studies elevates the threshold of excitability of LHRH neurons to norepinephrine (NE)_and/or pituitary responsiveness to LHRH, (3) preliminary MPN-ECS reduces thresholds of resposiveness of LHRH neurons to NE, and (4) the LHRH secreted after MPN-ECS sensitizes the pituitary gland to the subsequent small amounts of LHRH released following LC- or A1-stimulation.To provide answers to these questions, 3 experiments were performed in estrogen-treated OVX rats into which had been inserted the third ventricle and jugular cannulae. In the first study, the effects of artificial cerebrospinal fluid (ACSF) or ACSF + Ne (5,20, or 45 μg) on plasma LH concentrations in unanesthetized, unrestrained rats were examined. The intracerebroventricular (ICV) infusion of 5 μg of NE increased plasma LH by 61.3% above basal levels within 10 min whereas 20 or 45 μg NE elevated LH values 166.9 and 182.8%, respectively. The next study examined the effects of anesthetic drugs on LH response produced by ICV infusions of 45 μg of NE. Regardless of whether rats were anesthetized with ether, chloral hydrate, urethane, Saffan (alphaxolone + alphadolone) or ketamine + acepromazine, peak LH responses to ICV NE were not significantly different from unanesthetized controls. In a second study we observed that ICV NE (45 μg) markedly amplified and prolonged the release of LH after MPN-ECS. Moreover, the peak LH responses in these animals were approximately 10× greater than those obtained in rats which received ICV NE but not MPN-ECS. The third series of studies demonstrated that pituitary responsiveness to LHRH was not an important factor in dictating the LH response obtained after NE ICV infusions. These data suggest that LHRH neurons in estrogen-primed OVX rats are not particularily responsive to NE and that following MPN-ECS, LHRH neuronal responsiveness to this catecholamine markedly increases.     

Transforming growth factor-β1 exhibits delayed gene expression following focal cerebral ischemia

Transforming growth factor-β1 (TGF-β1) is a pleiotropic peptide growth factor. The expression of TGF-β1 mRNA in the focal ischemic cortex of rats was studied by means of Northern hybridization. A moderately low level of constitutively expressed TGF-β1 mRNA was detected following ohm surgery or in the contralateral (nonlschemic) cortex. A significant increase of TGF-β1 mRNA level in the ischemic cortex was observed at 2 days (3.2-fold increase compared to sham-operated animals, p < 0.01, N = 4) following permanent occlusion of the middle cerebral artery (PMCAO). The elevated TGF-μ1 mRNA expression was plateaued for up to 16 days (3–6-fold increase, p < 0.01) following PMCAO- This temporal profile for TGF-β1 mRNA expression in focal stroke was significantly delayed compared to that of TNF-α, IL-1β and IL-6 MRNA expressions as demonstrated previously which peaked at 12 h and decreased to almost basal levels by S days following PMCAO. Interestingly, the TGF-β1 mRNA expression profile was remarkably parallel with that of monocyte/macrophage accumulation in the ischemic cortex, as well as with the increased formation of extracellular matrix in the focal ischemic brain. These data suggest that TGF-β1 may play a role in anti-inflammatory process and in tissue remodeling following ischernic brain injury.     

Increase of interleukin-1β mRNA and protein in the spinal cord following experimental traumatic injury in the rat

Interleukin-1 beta (IL-1β) is a major mediator of inflammation and a growth promoter for many cell types that could play an important role in the consequences of traumatic spinal cord injury (SCI). In the present study, the expression of IL-1β and its mRNA was determined in the rat spinal cord following a standardized contusion injury. IL-1β mRNA, measured with quantitative RT-PCR, was significantly increased in the lesion site by 1 h after SCI (35.2±5.9 vs. 9.1±2.1 pg/mg RNA, n=3, P<0.05) and remained significantly higher than in the normal spinal cord for at least 72 h post-injury (p.i.). IL-1β mRNA levels in tissue immediately caudal to the lesion site did not change after the injury. IL-1β protein levels, measured by an ELISA, were determined at the lesion site and in cerebrospinal fluid (CSF) and serum samples. IL-1β levels in the CSF and serum were much lower than in the spinal cord. At the lesion site, IL-1β was increased significantly by 1 h p.i., peaked at 8 h (32.3±0.1 vs. 7.6±1.9, ng/g tissue, n=5, P<0.05) and remained significantly higher than normal through at least 7 days p.i. These results suggest that the increased IL-1β mRNA and protein levels are an early and local response at the lesion site that could trigger other, later, responses to traumatic SCI.