The steady-state kinetics of the butyrylcholinesterase-catalysed hydrolysis of butyrylthiocholine and thiophenyl acetate were shown to deviate from Michaelis-Menten kinetics. The ;best' empirical rate law was selected by fitting different rate equations to the experimental data by non-linear regression methods. The results were analysed in view of two alternative interpretations: (1) the reaction is catalysed by a mixture of enzymes, or (2) the activity is due to a single enzyme displaying deviations from Michaelis-Menten kinetics. It was concluded that the second alternative applies, and this conclusion was further supported by experiments involving simultaneous hydrolysis of alternative thiol ester substrates (butyrylthiocholine/thiophenyl acetate) as well as alternative thiol ester and oxygen ester substrates (butyrylthiocholine/phenyl acetate; thiophenyl acetate/butyrylcholine; acetylthiocholine/phenyl acetate). On the basis of the conclusion that a single enzyme is responsible for the activity, a molecular model is proposed. This model involves an acylated enzyme, and implies binding to the enzyme of one acyl group and one ester molecule, but not two ester molecules at the same time. Thus butyrylcholinesterase, which is structurally a tetramer, behaves functionally as a co-operative dimer, an interpretation in accordance with available data from active-site titrations.

Animals↗

PubMedDate not supplied

Interaction of drugs with the cyclic nucleotide generating enzymes in the central nervous system.

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Adenylyl Cyclase Inhibitors↗

PubMedDate not supplied

Chronic atropine treatment causes increase in VIP receptors in rat cerebral cortex.

Chronic atropine treatment (14 days, 20 mg X day-1 X kg-1 SC) caused a 75% increase in the number of VIP receptors in the rat cerebral cortex. The affinity of these receptors for 125I-VIP was not altered significantly by the atropine treatment. The same treatment led to a 20% decrease in VIP tissue levels. Muscarinic receptor number was also increased by 26%. The results indicate that interactions between VIP- and muscarinic receptors may be of importance in the rat cerebral cortex.

Animals↗

PubMedDate not supplied

VIP-sensitive adenylate cyclase, guanylate cyclase, muscarinic receptors, choline acetyltransferase and acetylcholinesterase, in brain tissue afflicted by Alzheimer's disease/senile dementia of the Alzheimer type.

Biochemical parameters were determined in autopsy material from several brain regions of thirteen patients with Alzheimer's disease/senile dementia of Alzheimer type (AD/SDAT) (mean age 75 years) and from brains of ten age-matched controls (mean age 76 years). Choline acetyltransferase specific activity was significantly lower in the nucleus caudatus, putamen, left thalamus, hippocampus and the cortex from gyrus hippocampus and the temporal lobe in AD/SDAT, acetylcholinesterase specific activity was significantly lower in the hippocampus, parietal and left frontal lobe in AD/SDAT samples than in corresponding samples from aged-matched controls. A compensatory increase of muscarinic receptors was found in the nucleus caudatus and left frontal lobe samples in AD/SDAT. Guanylate cyclase activity was not significantly altered in AD/SDAT in the brain regions examined. The basal, non-stimulated activity of adenylate cyclase was significantly (p less than 0.05) elevated in hippocampus samples from AD/SDAT patients and the enzyme activity stimulated by the vasoactive intestinal polypeptide VIP (2 microM) or forskolin (10 microM) was also elevated in AD/SDAT although not significantly.

Acetylcholinesterase↗

PubMedDate not supplied

Galanin and spinal nociceptive mechanisms: recent advances and therapeutic implications.

Galanin is a peptide consisting of 29 or 30 (in humans) amino acids that is present in sensory and spinal dorsal horn neurons. Endogenous galanin may have an important modulatory function on nociceptive input at the spinal level. In addition, exogenously administered galanin exerts complex effects on spinal nociceptive transmission, where inhibitory action appears to predominate. Peripheral nerve injury and inflammation, conditions associated with chronic pain, upregulate the synthesis of galanin in sensory neurons and spinal cord neurons, respectively. Hence, the sensory effect of galanin may be increased under these conditions, raising the possibility that modulation of the activity of the galanin system may produce antinociception.

Analgesics↗

PubMedDate not supplied

Simultaneous measurement of brain and core temperature in the rat during fever, hyperthermia, hypothermia and sleep.

The neuropathological outcome of metabolic, vascular or mechanical insults to the CNS depends on brain temperature; mild hypothermia is neuroprotective, whereas elevated brain temperature can cause additional neural damage. Studies in both animals and humans have shown that the core and the brain temperature do not always concur with one another. It is therefore important to develop methods for monitoring brain temperature. This paper describes an animal model (the rat) in which we have developed a method to measure, at thermoneutral ambient temperature, the brain and core temperature concomitantly, during different drug treatments. We have used this animal model to study body temperature during fever (induced by human recombinant IL-1 beta, 5 microgram/kg, i.p.), stress-induced hyperthermia (handling of the animal), hypothermia (induced by (+/-)-8-hydroxy-2-dipropylaminotetralin hydrobromide, 0.5 mg/kg, i.p. ) and sleep (non-induced, other than by light and diurnal variation). We show that the thermal curves are similar in the brain and the peritoneum, independent of the thermal state.

8-Hydroxy-2-(di-n-propylamino)tetralin↗

PubMedDate not supplied

Autoradiographic quantitation and anatomical mapping of 125I-galanin binding sites in the rat central nervous system.

The distribution of 125I-galanin binding sites in the rat central nervous system was studied by the use of autoradiography, and the amount of peptide bound was determined by the use of microdensitometry. The amount of 125I-galanin bound in various CNS regions ranged from non detectable to 0.40 pmol per gramme tissue at a concentration of 1.5 nM 125I-galanin. The anatomical mapping revealed high density of binding sites in the telencephalon, where labelling was seen in the entorhinal, perirhinal and piriform cortices, in most septal nuclei, in the amygdala and in the hippocampal formation. In the diencephalon binding sites were densely accumulated in the hypothalamus, including the median eminence and periventricular nucleus as well as in the dorsal and medial thalamic nuclei. In the mesencephalon binding sites were seen in many nuclei including the dorsal raphe nucleus, the periaqueductal central grey and pars compacta of the substantia nigra. The pons/medulla revealed high density of binding sites in the locus coeruleus, parabrachial nuclei and in the dorsal vagal complex, while binding in the spinal cord was seen in laminae I, II and X as well as in the intermediolateral horn. The distribution of 125I-galanin binding sites corresponded well to the localization of the peptide as revealed in several earlier immunohistochemical and radioimmunoassay studies, as well as to several previously reported effects of galanin on central autonomic functions.

Animals↗

PubMedDate not supplied

Acute and chronic studies on functional aspects of coexistence.

Autoinhibition of acetylcholine release by the coexisting peptide galanin in the septal afferents to the hippocampus of the rat was examined in tissue slices from the hippocampus. Galanin inhibits the evoked release of the coexisting neurotransmitter, acetylcholine, in the ventral hippocampus, providing an example of autoinhibition of release of a neurotransmitter by one of the coexisting neurotransmitters. The galanin mediated inhibition of the acetylcholine release is a complement to the well known strong cholinergic autoinhibition. The effects of the coexisting galanin and acetylcholine on several second messenger systems were also examined: acetylcholine acting at muscarinic receptors depresses cyclic adenosine 3',5'-monophosphate and stimulates elevation of cyclic guanosine 3',5'-monophosphate levels, whereas neither cyclic adenosine 3',5'-monophosphate nor cyclic guanosine 3',5'-monophosphate levels were affected by galanin (1 microM). Galanin however inhibited partly the muscarinic stimulation of phosphoinositide breakdown, suggesting that inositol phosphate(s) or diacylglycerol may act as second messenger(s) of the galanin action in the hippocampus. The effects of chronic changes in firing rate on the coexisting neurotransmitters in the rat ventral spinal cord containing serotonin, thyrotropin releasing hormone, substance P and substance K were examined. The tissue levels of the coexisting transmitters were studied in rats chronically treated with imipramine (14 days; 2 x 10 mumoles/kg/day) and zimelidine (14 days; 2 x 10 mumoles/kg/day). Upon treatment with zimelidine the tissue levels of the serotonin metabolite 5-hydroxyindoleacetic acid fall by 32% while thyrotropin releasing hormone levels seem to increase 35% and substance P/substance K levels also increase 48 and 72% respectively. Imipramine treatment resulted in similar although less pronounced changes.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗

PubMedDate not supplied

Interleukin-1 stimulates the expression of type I and type II interleukin-1 receptors in the rat insulinoma cell line Rinm5F; sequencing a rat type II interleukin-1 receptor cDNA.

The insulin secreting rat Rinm5F cells are often used to study the cytotoxic actions of interleukin-1 (IL-1) on pancreatic beta-cells. We demonstrate here that Rinm5F insulinoma cells express both type I and type II interleukin-1 receptor (IL-1R) mRNAs and gene products. IL-1R agonists, recombinant murine IL-1 alpha (rmIL-1 alpha, 10 ng/ml) and recombinant rat IL-1 beta (rrIL-1 beta, 100 pg/ml or 10 ng/ml) induce the upregulation of mRNA expression for both types of IL-1 receptors (IL-1Rs). This effect of rrIL-1 beta is antagonised by preincubation with recombinant human interleukin 1 receptor antagonist protein (rhIL-1ra, 5 micrograms/ml). Furthermore, this rrIL-1 beta induced upregulation of IL-1R mRNAs is blocked by actinomycin D (7.5 micrograms/ml), whereas cycloheximide (20 micrograms/ml) has no effect. The phorbol ester PMA (20 nM) upregulates the expression of mRNAs both IL-1 receptors, whereas glucose (50 mM) upregulates the expression of the type I IL-1R mRNA only. Pretreatment of cells with pertussis toxin (100 ng/ml) partially blocks the rrIL-1 beta induced expression of mRNA for the type I and, to a lesser extent, the type II IL-1R. Incubation of the cells with rrIL-1 beta also induces a time-dependent expression of c-fos, interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF-alpha) mRNAs. Binding studies with 125I-recombinant human IL-1 beta (125I-rhIL-1 beta) indicate that IL-1R gene products, with the ligand binding characteristics of the type I IL-1R, are constitutively present on Rinm5F cells. Treatment with rrIL-1 beta (6h) increases the number of 125I-rhIL-1 beta binding sites on Rinm5F cells. We have also demonstrated that the number of type II IL-1R binding sites increases after induction with rrIL-1 beta (6h), by indirect immunofluorescence using a monoclonal antibody (ALVA 42) raised against the human type II IL-1R. Furthermore, we have sequenced the type II IL-1R cDNA in the rat insulinoma Rinm5F cells. The comparison of the amino acid sequence of the rat type II IL-1R with that of the mouse and human type II IL-1Rs shows 90% and 62% amino acid identity, respectively. The most important difference between the human and murine type II IL-1Rs, and this rat type II IL-1R cDNA, is an open reading frame coding for a six amino acid longer, strongly charged (QIKEMK), cytosolic domain.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗