Central pyrogenic activity of muramyl dipeptide.

Fever can be elicited in the rabbit by the intravenous administration of relatively large doses of a synthetic immunoadjuvant, N-acetylmuramyl-L-alanyl-D-isoglutamine, or muramyl dipeptide (MDP). This response could be mediated by endogenous pyrogen because MDP has been shown to induce their production both in vivo and in vitro. The results reported here show that intracisternal injection of minute amounts of MDP could elevate fever without activating the release of endogenous pyrogen in the plasma or in the cerebrospinal fluid. Moreover, indomethacin inhibited hyperthermia produced by intracerebroventricular administration of MDP. Therefore, our findings argue in favor of a direct effect of the glycopeptide on the thermoregulatory centers besides its indirect effect through the production of leukocytic pyrogen. This molecule apparently represents the minimal requirement for the pyrogenicity of bacterial peptidoglycan because administration, even by the intracerebral route, of a mixture of muramic acid and of its dipeptide moiety did not elicit fever.

Photoelectron spectroscopy characterization of diamond-like carbon films.

The electronic states of diamond-like hydrogenated carbon (DLC) films were studied by synchrotron radiation photoelectron spectroscopy. The valence band spectra measured at different excitation energies show the gradual emergence of the p-pi band in relation to the sample annealing and ion bombardment amorphization. The p-pi band of the annealed DLC was characterized by localized p(z) states, while the formation of the amorphous carbon surface was accompanied by appearance of the delocalized p(z) states, which reduce the optical gap. A simple approach permitting the extraction of the 2p band shape from the photoelectron spectra is proposed.

Prenatal dihydrotestosterone differentially masculinizes tonic and surge modes of luteinizing hormone secretion in sheep.

The control of LH secretion in sheep is sexually differentiated. Males begin to reduce their sensitivity to inhibitory steroid feedback, leading to a pubertal increase in tonic LH secretion by 10 weeks of age, but females remain hypersensitive until 30 weeks. Moreover, only females can respond to the positive feedback action of estradiol to produce a preovulatory LH surge. Prenatal exposure of the female lamb to testosterone masculinizes tonic LH and abolishes the LH surge postnatally. However, the type of steroid involved is not known because testosterone can be converted to estradiol or dihydrotestosterone (DHT). This study tested the hypothesis that DHT, which cannot be converted to an estrogen, masculinizes tonic LH without defeminizing the LH surge. Pregnant ewes were treated with DHT (800, 400, or 200 mg/week) during the critical period for sexual differentiation of gonadotropin secretion (days 30-90; 145 days is term). To evaluate the time of the decrease in responsiveness to steroid inhibition, a constant steroid feedback signal was produced. At 4 weeks of age, androgenized females (800 mg, n = 5; 400 mg, n = 4; 200 mg, n = 5) and control males (n = 7) and females (n = 9) were gonadectomized and implanted with a SILASTIC brand estradiol capsule. Tonic LH secretion in males began to increase at 6.7 +/- 0.5 weeks (mean +/- SEM). In DHT-treated females, the LH increase began at the same time (800 mg DHT, 10.7 +/- 3.9 weeks; 400 mg DHT, 9.9 +/- 5.9 weeks; 200 mg DHT, 7.1 +/- 4.9 weeks). This was several months earlier than in control females (29.1 +/- 0.8 weeks; P < 0.05). After puberty, estradiol induced LH surges in 8 of 9 control females and 11 of 12 DHT-treated females, but not in any control males. These results lead to the hypothesis that in the sheep, distinct requirements exist for differentiation of 2 types of reproductive hormone control systems, and that conversion of testosterone to an estrogen is not essential for both. Aromatization is necessary to prevent the surge control of GnRH from operating in the male, but nonaromatizable androgens differentiate the tonic control to permit high GnRH secretion earlier in life.

A modulating role of prostaglandins in contractions of the guinea-pig ileum.

1 The role of prostaglandins in contractions of the guinea-pig ileum evoked either directly by acetylcholine or indirectly by angiotensin and by coaxial stimulation has been investigated.2 Prostaglandin E(2) in low concentration (6 nM) slightly augmented both types of contraction. Indomethacin, an inhibitor of prostaglandin synthesis, markedly reduced the indirectly evoked contractions but did not affect contractions in response to acetylcholine. The addition of prostaglandin E(2) to the preparation treated previously with indomethacin restored the effect of indirect stimulation.3 The pretreatment of the preparation with guanethidine or alpha-methyl-p-tyrosine prevented the inhibitory effect of indomethacin on indirectly evoked contractions. Prostaglandin E(2) addition to such preparations considerably augmented both types of contraction.4 The stimulation of non-cholinergic, non-adrenergic inhibitory nerves in the taenia coli and ileum preparations evoked hyperpolarization and relaxation of the preparations followed by action potentials and contraction. These responses were not changed by indomethacin pretreatment and prostaglandin E(2), but rebound contraction was sometimes augmented by the prostaglandin.5 Two mechanisms for the effects of prostaglandin E(2) are suggested: a direct effect on smooth muscle, and an indirect action through the sympathetic nerves which by release of noradrenaline affect the acetylcholine release from parasympathetic nerve endings.

The topography of cholinergic transmission in the mechanism of drug action at muscarinic synapses of the guinea-pig ileum.

The pharmacology of cholinergic neurogenic responses evoked by the participation of only the endings of axon terminals was compared to that of responses evoked by participation of the more proximal parts of the terminals also. Myenteric plexus-longitudinal muscle strips of the guinea-pig ileum were drawn through narrow orifices in 2 rubber membranes dividing a bath into 3 separate compartments. Oral segments were stimulated electrically by single impulses or by trains and local neurogenic contractions were evoked. The contractions of the aboral segment due to nerve impulses transmitted from the oral segment via the middle segment were also recorded. The opioid ligands ketocyclazocine and [D-Ala2,MePhe4,Met(O)5-ol]enkephalin and noradrenaline inhibited the twitches of the aboral segment evoked by oral segment stimulation more than the local twitches of the oral segment when these agents were applied directly to the respective compartments. The twitches of the aboral segment were also inhibited by the application of these drugs into the middle compartment adjusted to 10 mm width. Verapamil and the alkaline earth metal ions cobalt and lanthanum had similar effects. 4-Aminopyridine increased twitch amplitude more in the aboral segment than in the oral segment when applied directly; similar effects in the aboral segment were seen when the agents were applied to the middle compartment. The action of atropine, papaverine, d-tubocurarine and prostigmine did not discriminate between twitches in the oral and aboral segment when applied directly and all drugs except prostigmine were without effect when applied to the middle compartment.(ABSTRACT TRUNCATED AT 250 WORDS)

Post-tetanic potentiation in the innervated smooth muscle preparation of the guinea-pig ileum.

The phenomenon of posttetanic potentiation of electrically evoked twitch contractions in the myenteric plexus-longitudinal muscle of the guinea-pig ileum was observed when (1) post-tetanic inhibition had been prevented by naloxone, an opiate receptor antagonist, and (2) the twitches had been diminished by indomethacin, an inhibitor of prostaglandin synthesis. The origin of posttetanic potentiation at this muscarinic synapse appeared to be presynaptic.

The analysis of post-tetanic potentiation in guinea-pig ileum longitudinal muscle strip.

The effect of tetanus on the twitch responses of the longitudinal muscle-myenteric plexus preparation of the guinea-pig ileum to electrical stimuli was investigated in the presence of naloxone and indomethacin. Naloxone was used to prevent post-tetanic twitch inhibition due to the release of endogenous opiate ligands, and indomethacin to diminish pretetanic twitch height. Twitch contractions following tetanus were potentiated in the presence of both drugs. The optimal stimulation parameters for the manifestation of post-tetanic potentiation (PTP) were determined; tetanic stimulation: 30 Hz for 25 s. supramaximal impulse intensity; twitches: 0.04 Hz, low impulse intensity. PTP was also obtained when indomethacin was replaced by noradrenaline or adenosine, i.e. by drugs whose mechanism of action also includes a presynaptic effect. Postsynaptic changes in contractility cannot fully account for the observed PTP of twitches judging from the smaller effect of tetanus on acetylcholine-evoked contractions. The hypothesis of a presynaptic origin of PTP at this muscarinic synapse was corroborated in the experiments where acetylcholine stores were labelled with [3H] choline and the release of the label was increased during PTP. Furthermore, a bioassay showed that the output of endogenous acetylcholine in the post-tetanic interval was increased in the presence of naloxone plus indomethacin but not in their absence. The fact that PTP was also observed in the absence of any drug, if the tetanic stimulation was short (10 s) and of low impulse intensity, suggested its possible physiological significance.

Post-tetanic potentiation at the nerve-muscle junction in the longitudinal muscle of the guinea-pig ileum. Possible role of substance P.

The effect of short tetanic stimulation (30 Hz for 25 s) on the following twitch responses of the myenteric plexus-longitudinal muscle preparation of guinea-pig ileum to electric stimulation (0.1 Hz) was investigated in the presence of naloxone and indomethacin. Post-tetanic potentiation (PTP) of the twitches observed in control experiments was abolished in preparations desensitized by substance P but it was not affected in preparations desensitized by serotonin or pretreated with methysergide. Immediately after 5 min tetanic stimulation a decreased sensitivity to substance P but unchanged sensitivity to serotonin were observed. Electromyogram (EMG) of the longitudinal muscle layer was picked up 4 and 10 mm aborally from the stimulation site in response to 1 to 16 impulse trains delivered at 100 Hz. In control conditions only the longer trains triggered this neurogenic response at the distal recording site. In the presence of substance P but not serotonin facilitation occurred so that the distal site was frequently recruited to respond with an EMG even to single impulses. A substance P-like compound rather than serotonin may be a candidate for the neuromodulator or neurotransmitter substance involved in PTP and changes in the response topography of muscarinic transmission.

Mutual connections of the raphe system and hypothalamus in relation to fever.

Previous studies in our laboratory have shown that lesions in certain brain stem regions prevent elevations of body temperature after administration of bacterial pyrogen. In the present experiments wer examined the morphology of the connections of these "brain stem thermoregulatory centers" which are represented in all raphe and paraphe nuclei of the brain stem reticular formation. The methods of anterograde degeneration and tracing of retrograde transport of horseradish peroxidase were used to identify afferent and efferent connections within this "thermoregulatory field." Abundant mutual connections between the raphe nuclei and hypothalamus were found. All nuclei of the raphe system receive afferents from the medial and lateral hypothalamus. All raphe nuclei have efferent projections to the medial reticular formation, and the raphe nuclei of the pons and mesencephalon provide ascending fibers to the hypothalamus. A lesion of any part (origin, course, termination field) of this mutual raphe-hypothalamic pathway system will prevent development of fever in response to bacterial pyrogen.

The involvement of brain structures in the adjuvant effect of muramyl dipeptide.

With the aid of small electrolytic lesions we have studied the possible participation of brainstem structures in the adjuvant activity of muramyl dipeptide (MDP). The results suggest the involvement of the serotonergic groups B6.7.8 and serotonergic pathways in the upper region of the reticular formation. Since lesions in the caudal parts of reticular formation in the area of aminergic groups A1.3.5.7 with the participation of corresponding pathways also influenced the adjuvant effect of MDP the possible role of noradrenaline is also implicated.

Interactions between the duration of stimulation and noradrenaline on cholinergic transmission in the myenteric plexus-smooth muscle preparation.

Output of acetylcholine (ACh), electromyogram (EMG) recordings and contractions of myenteric plexus-longitudinal muscle strip preparations from the guinea-pig ileum were studied during stimulation by single impulses or by trains (30 Hz; 2 to 128 impulses) under control conditions and in the presence of noradrenaline (NA). During supramaximal stimulation NA (2.5 microM) inhibited both contractions of the smooth muscle and the release of ACh evoked by single impulses more effectively than those evoked by train stimulation so that in a train of 4 impulses the output of ACh per impulse after the 2nd to 4th impulses was 69 to 104% higher than the output after the 1st impulse. During submaximal stimulation, contractions and ACh release evoked by single impulses were almost completely inhibited by NA. The neurogenic EMG, a direct consequence of the localized action of released transmitter (ACh), was recorded in the longitudinal muscle 4 and 10 mm aborally from the focal stimulation site. The incidence of the neurogenic response was much higher at the proximal (4 mm) than at the distal (10 mm) site and was proportional to the number of impulses in a train (100 Hz). NA inhibited propagation of the neurogenic response evoked by single impulses whereas its effect during train stimulation was less. It is concluded that in the course of train stimulation, sites of transmission more distant from the stimulation focus was recruited, and consequently the secretion of ACh in succeeding impulses was enhanced. NA could interfere with this process; it might inhibit the invasion by action potentials of cholinergic nerve terminal varicosities, thereby reducing the release of ACh.

Neuroendocrine immune interactions in health and disease.

The purpose of this paper is to review ways in which the neurohormonal system can interact with the immune system and to outline the main mechanisms which are involved in this interaction. Experimental as well as clinical evidence is presented to support the existence of a close interaction and bi-directional communication between the central nervous and immune systems. The role of major endocrine mechanisms and hormones is discussed. The evidences from experimental work to support the roles of the nervous system with neurotransmitters, the endocrine system with hormones, and the immune system with cytokines are presented. Aging, depression and cancer have a high degree of co-association and share mechanisms which result in cellular immune deficiency. Hormone therapy, zinc replenishment, antidepressants, immunomodulators like MDP act on these pathways to upregulate and improve cellular immunity. The authors believe that the central nervous system (CNS)-immune interaction is an important new frontier to be considered for new combination therapy in diseases with cellular immune deficiency such as cancer particularly in the aged with depression.

Stimulation of nonspecific immunity, haemopoiesis and protection of mice against radiation injury by 1-adamantylamide-L-alanyl-D-isoglutamine incorporated in liposomes.

1-Adamantylamide-L-alanyl-D-isoglutamine (adamantylamide dipeptide (AdDP)) belongs to a group of desmuramyl muramyl peptide derivatives which are able to protect an organism from some viral infections. Encapsulation of AdDP to egg phosphatidyl choline liposomes and the targeting of this drug to lymphatic node macrophages via subcutaneous (s.c.) administration proved to be the efficient way to protect mice against irradiation when administered s.c., 24 h prior to lethal gamma-irradiation (long-term survival rate in the range of 40% compared with 0% in saline or free drug control). Parameters characteristic for the recovery of haemopoiesis in the bone marrow (number of granulocyte-macrophage haemopoietic progenitor cells, granulocyte-macrophage colony forming cells (GM-CFC)) were significantly improved in comparison with the controls and free drug on day 10 after 6.5 Gy irradiation. The haemopoietic effect was observed in the broad application time window (72 h before and 48 h after irradiation). Very high radioprotective effect of s.c. administered liposomal AdDP (L-AdDP) can be explained (together with induction of haemopoiesis) by the effective and long-lasting activation of nonspecific immunity, which withholds the onset of septicemia in early days after irradiation. Induction of nonspecific immunity was proven in Candida albicans infectious model. L-AdDP significantly increased both the survival time and score (about 40% survival compared with 0% in controls and free drug). In conclusion, L-AdDP could be therapeutically beneficial to moderate the haemopoietic damage (undesirable effect of radiotherapy or chemotherapy) and induce the non-specific immunity to support the antimicrobial treatment of immunocompromised patients.

Past, present and future of psychoneuroimmunology.

Psychoneuroimmunology was for the first time comprehensively described about 20 years ago. The influence of mental status on the course and outcome of a number of diseases, however, was suspected a long time before. Also the links between mental affective disorders and the immune status were repeatedly suggested. The authors in this paper shortly reviewed the most important clinical as well as experimental evidence which at present strongly supports the concept of a close and bidirectional communication between central nervous, neuroendocrine and immune systems. The most important anatomical, physiological as well as pharmacological experimental data, which were obtained by the authors during 20 years of research in this field, are presented. The data strongly suggest that in the very next future we will not only better understand a very complex communication between mind and body, but also completely new types of compounds might become available.

Potential role of cannabinoids in Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disorder caused by a progressive loss of dopaminergic neurons of the substantia nigra, resulting from an oxidative stress. The lack of dopaminergic neurons is reflected by a disturbed balance of the neural circuitry in the basal ganglia. Cannabinoids might alleviate some parkinsonian symptoms by their remarkable receptor-mediated modulatory action in the basal ganglia output nuclei. Moreover, it was recently observed that some cannabinoids are potent antioxidants that can protect neurons from death even without cannabinoid receptor activation. It seems that cannabinoids could delay or even stop progressive degeneration of brain dopaminergic systems, a process for which there is presently no prevention. In combination with currently used drugs, cannabinoids might represent, qualitatively, a new approach to the treatment of PD, making it more effective.

Drug effects on attack defense and escape in mice.

The relative specificity and potency of action of 32 neuropsychotropic drugs was assessed on attacks, defensive upright postures and escapes occurring in singly-housed male mice during interactions with non-aggressive strange males. Scopolamine was most potent in reducing attacks while apomorphine was most active in stimulating attacks. Defenses and escapes were inhibited most efficiently by pentobarbital and diazepam, while L-tryptophan was most active of the tested drugs in stimulating defenses and escapes. Aminooxyacetic acid and valproate inhibited both attack and defensive-escape behavior at relatively low doses. Inhibition of attacks by many drugs tested could be explained by anticholinergic, serotonergic or gabaergic effects while stimulation of attacks may be due to dopaminergic effects. Gabaergic drug actions seem to inhibit while serotonergic effects might stimulate defenses and escapes.

Possible role of prostaglandins in post-tetanic potentiation at the nerve-muscle junction in the longitudinal muscle strip of guinea-pig ileum.

The effect of tetanic stimulation on the twitch responses of the longitudinal muscle-myenteric plexus preparation of the guinea-pig ileum to electrical stimulation was investigated in the presence of naloxone. Under this condition, or after the addition of PGE2, twitch contractions were maximal and no potentiation of twitches following tetanus was observed. In the presence of indomethacin (1 mumol litre-1) twitches were diminished and post-tetanic potentiation (PTP) was manifested. PTP was seen with indomethacin concentrations of 1 to 20 mumol litre-1 or after simultaneous addition of diphloretin phosphate (16 mumol litre-1). Thus it seems unlikely that the effect of prostaglandins released during tetanic stimulation would be of key importance for the manifestation of PTP. Rather it is thought that a decrease in the release of acetylcholine from motor nerve terminals, and consequently smaller twitches in the presence of indomethacin, offer favourable conditions for PTP.

Non-synaptic cholinergic modulation of neurogenic twitches of the guinea-pig ileum.

The effect of cholinergic and anticholinergic compounds on conduction of neuronal excitation has been studied in myenteric plexus-longitudinal muscle strips from the guinea-pig ileum. A preparation in a special triple bath was drawn through two rubber membranes dividing the strip into three segments. Neurogenic stimulation of the oral segment set up nerve action potentials propagating aborally across the middle segment (10 mm) so that the aboral segment might be also invaded, eventually. Drugs were added to the middle segment to affect neuronal propagation (non-synaptic effects) which was monitored by twitch height of the aboral segment. The application of acetylcholine to the middle segment augmented aboral twitches. The effects of nicotine, pilocarpine and oxotremorine were selectively blocked by (+)-tubocurarine, pirenzepine and atropine, respectively. The effect of acetylcholine was suppressed by pirenzepine and atropine and mimicked by doubling of KCl concentration. The effect of acetylcholine may be thus explained by the facilitated propagation of nerve action potentials in partially depolarized cholinergic terminals via stimulation of muscarinic receptors. The adenylate cyclase system is not directly involved in the mechanism of muscarinic facilitation of neuronal propagation in the terminals; however, it may participate in the modulation of a final common effector mechanism.

Increased aggressiveness and lower brain serotonin levels in offspring of mice given alcohol during gestation.

Male adult offspring of mice given alcohol during gestation showed more aggressive and locomotor activities and had lower brain serotonin concentrations than did control offspring.

Non-junctional modulation of neurogenic twitches of the guinea-pig ileum by some peptides and other compounds in the triple bath.

The effects of some neuropeptide transmitter candidates and of some other neurotoxins or drugs on conduction of neural excitation were studied in myenteric plexus-longitudinal muscle strips from the guinea-pig ileum. A preparation in a special triple bath was drawn through two rubber membranes dividing the strip into three segments. Neurogenic stimulation of the oral segment set up nerve action potentials propagating aborally across the middle segment so that the aboral segment might also be invaded. Drugs were added to the middle segment to affect neuronal propagation (non-junctional effects) which was monitored by twitch amplitude of the aboral segment. The application of bradykinin and cromakalim did not affect aboral twitches although strong contractile and relaxatory effects were observed when the drugs were applied directly to the aboral segment; no neurogenic effects thus manifested. Capsaicin and neurotensin, when applied both to the middle and aboral segments, elevated the tone of the preparations accompanied with a decrease in twitch amplitude; these effects may have been due to neurogenic stimulation and release of other motor neurotransmitters. The application of VIP, apamin and dendrotoxin to the middle as well as to the aboral segments augmented aboral twitches, which might be at least partly due to facilitation of nerve action potential propagation in nerve terminals of cholinergic motor fibres.