I.M. Gardiner

Clinical and functional investigation of 10 missense mutations and a novel frameshift insertion mutation of the gene for copper-zinc superoxide dismutase in UK families with amyotrophic lateral sclerosis

Article abstract-Mutations of the gene SOD-1, which encodes the enzyme copper-zinc superoxide dismutase, occur in patients with a familial form of amyotrophic lateral sclerosis (ALS). We investigated 71 families with more than one individual affected by ALS for clinical features and SOD-1 mutations. Mutations were identified in 14 families, indicating the presence of SOD-1 mutations in around 20% of this population. There were 10 different heterozygote missense point mutations in eight different codons, and a novel two-base frameshift insertion (132insTT), which leads to substitution of aspartic acid for glutamic acid at codon 132, and a premature stop codon at 133, with predicted truncation of the protein. SOD enzyme activity was reduced to around 50% of normal in individuals with SOD-1 mutations, and may be a useful predictor for the presence of these mutations. A predilection for disease onset in the lower limbs appears to be a distinguishing feature of familial ALS with SOD-1 mutations, and accords with findings in transgenic mouse models. In general, the finding of an SOD-1 mutation does not accurately predict a prognosis or disease severity. NEUROLOGY 1997;48: 746-751

Stimulatory effect of N-methyl aspartate on locomotor activity and transmitter release from rat nucleus accumbens

N-Methyl-aspartate (NMA), an agonist at central glutamate receptors, elicited prolonged and intense locomotor activity when injected into the nucleus accumbens septi (NAS) in subconvulsive doses (3-10 micrograms bilaterally). This effect was antagonised by intra-accumbens injection of the specific NMA antagonist, aminophosphonovaleric acid (APV) in a dose (3.0 micrograms bilaterally) that was without intrinsic effect when given on its own. Intra-accumbens injection of APV also suppressed locomotor hyperactivity elicited by intra-accumbens injection of DA (50 micrograms bilaterally) in rats pretreated with nialamide. In vitro release of [3H]-acetylcholine in accumbens tissue slices was significantly increased in the presence of NMA (30 microM) or N-methyl-D-aspartate (NMDA) (15 microM). Both effects were antagonised by APV (30 microM). Similar results were obtained with tissue slices of rat corpus striatum. These results suggest that locomotor stimulation by intra-accumbens NMA is mediated by an action on the mesolimbic dopaminergic neuron, either directly or via a cholinergic interneuron. In addition, activity at the glutamate synapse may be enhanced by the presence of DA affecting glutamate release and/or reuptake.

Analysis of muscarinic receptor concentration and subtypes following lesion of rat substantia innominata.

Cholinergic neurones located in the nucleus basalis of Meynert (NBM) in the substantia innominata (SI) of primates are known to project to cerebral cortex and cell loss in NBM is thought to be associated with the cholinergic deficit seen in Alzheimers disease. We have examined in rats the effect of lesion of SI with kainate (1 microgram/0.5 microliter) on acetylcholine esterase (AChE) activity, muscarinic receptor number and subtypes in cerebral cortex at 1, 2 and 4 weeks. The area of lesion was assessed histologically. AChE activity was significantly reduced in frontal and parietal cortex ipsilateral to the lesion compared to the contralateral side by 37 and 30%, respectively, at 1 week. The reduction in parietal cortex at 4 weeks (16%) was significantly attenuated. Muscarinic receptor number was reduced in cerebral cortex ipsilateral to the lesion at the 3 time periods measured, being reduced by 14 and 17% in the frontal and parietal cortex, respectively, at 1 week. Changes in receptor number and AChE activity correlated with the size of lesion. Low affinity agonist binding sites and high affinity pirenzepine binding sites were also analyzed and found to be significantly reduced by lesion of SI. The proportions of high and low affinity agonist binding sites and subtypes of pirenzepine binding sites were, however, not significantly affected by lesion.

Action of apomorphine, bromocriptine and lergotrile onγ-aminobutyric acid and acetylcholine release in nucleus accumbens and corpus striatum

The effect of three dopamine agonists, apomorphine, bromocriptine and lergotrile, was tested on the release ofγ-aminobutyric acid, (GABA) and acetylcholine (ACh) from tissue slices of rat nucleus accumbens and striatum. All three agentsin vitro caused a dose dependent depression of the K+-evoked release of [14C]-GABA in corpus striatum. This effect was also obtained followingin vivo drug application and when endogenous GABA release was determined. A similar depression of GABA release was obtained in the nucleus accumbens. Both dopamine and dibutyryl adenosine-3′∶5′-cyclic monophosphoric acid inhibited the K+-evoked release of [14C]-GABA in corpus striatum. This inhibitory effect was not reversed by sulpiride. Bromocriptine and lergotrile also depressed the K+-evoked release of [3H]-acetylcholine from tissue slices of corpus striatum but not nucleus accumbens, as has previously been demonstrated for dopamine and apomorphine. In contrast, sulpiride enhanced the release of [3H]-acetylcholine and molindone reversed the apomorphine inhibition of [3H]-acetylcholine release. These results indicate that dopaminergic agents may influence the release of both GABA and ACh in the corpus striatum but only GABA in the nucleus accumbens.

Inhibitory effect of 1,2,3,4-tetrahydro-9-aminoacridine on the depolarization-induced release of GABA from cerebral cortex

1,2,3,4‐Tetrahydro‐9‐aminoacridine (THA) has an inhibitory effect on the activity of acetylcholinesterase which has led to its use in the treatment of Alzheimers disease. Other actions of THA include the inhibition of voltage‐dependent ion channels. In this paper we describe the effect of THA on the depolarization‐induced release of [14C]‐γ‐aminobutyric acid (GABA) from tissue slices of rat cerebral cortex. THA produced a dose‐dependent inhibition of the 30mM K+‐evoked release of [14C]‐GABA with an IC50 of 56 μm. The maximal response was an 84% inhibition of the evoked response. THA (up to a concentration of 1 mm) had no effect on the basal release of GABA. A similar inhibitory effect on the K+‐evoked release of [14C]‐GABA was seen with 4‐aminopyridine (4‐AP) but no inhibition was obtained with tetraethylammonium up to a concentration of 20 mM. The maximal inhibitory effect of 4‐AP (39%) occurred at 1mM (IC50 of 112 μm) and this response was much smaller in magnitude than that obtained with THA.

Muscarinic receptors discriminated by pirenzepine are involved in the regulation of neurotransmitter release in rat nucleus accumbens

1 The effect of pirenzepine, a selective muscarinic antagonist, was tested on the oxotremorine facilitation of the K+‐evoked release of [14C]‐dopamine from tissue slices of rat nucleus accumbens. 2 The effect of pirenzepine was compared with that of scopolamine and other antagonists which show no heterogeneity in their action on muscarinic receptors in order to determine whether a selective action at a single receptor subtype, M1 or M2, could be distinguished. 3 Pirenzepine and scopolamine both antagonized the oxotremorine‐induced (EC50 = 3 × 10−7M) facilitation of [14C]‐dopamine release with pA2 values of 7.5 and 8.9 respectively. This result indicated that the high affinity pirenzepine receptor (M1) was involved in this response. Low concentrations of 3‐quinuclidinyl benzilate (3 × 10−10 M), N‐methylscopolamine (3 × 10−9M) and methyl atropine (10−8 M) also abolished this facilitatory effect of oxotremorine.

A Study of Adaptive Responses in Cell Signaling in Migraine and Cluster Headache: Correlations Between Headache Type and Changes in Gene Expression

The project was an investigation into whether changes in the expression of G-proteins underlie altered cell signaling in migraine and cluster headache. The basis for this assumption is that altered physiological responses are seen in migraineurs and that differences in cell signaling are detected biochemically in various cell types isolated from peripheral blood. Levels of three G-protein mRNAs—Gsα, Giα, and Gα, were quantified in lymphocytes from clinically well-defined migraine and cluster headache patients and correlated with headache type and influence of drug treatment. Giα mRNA was reduced by 50% in all migraine patients compared with control subjects; similarly in patients with or without aura, in patients with a migraine headache at the time of sampling, and patients in a quiescent state. No reduction in the levels of Gsα or Gα mRNA were seen in migraine patients. A smaller reduction was seen in cluster headache patients, most marked in those without medication. Levels of Gsα mRNA were significantly reduced in cluster headache patients compared with migraine patients. The marked down-regulation of Giα mRNA in migraine, whether quiescent or acute, indicates either an adaptive response to headache in this group of patients or that low levels of Giα mRNA make individuals more susceptible to migraine.

Pharmacological analysis of red-wine-induced migrainous headaches

Abstract We describe a series of experiments designed to investigate the mechanisms by which headaches can be triggered by red wine in a small minority of migraine patients. Some red wines are particularly potent releasers of serotonin from platelet stores, but these are no more effective as triggers of headache in sensitive patients. Both the selective 5-HT2A antagonist ketanserin and the non-selective 5-HT2ABC antagonist pizotifen blocked the majority of headaches, and we then thought the antihistamine properties of these two drugs might be important. In a third experiment, however, the H1 antagonist mepyramine did not convincingly antagonise the response to red wine. Plasma levels of the enzyme diamine oxidase, which metabolises histamine, were lower in all migraine patients, whether or not they were sensitive to red wine. The results reported here do not permit making firm conclusions; nevertheless it seems that different pharmacological receptors may be responsible in different patients.

Excitotoxin induction of ornithine decarboxylase in cerebral cortex is reduced by phospholipase A2 inhibition

The enzyme ornithine decarboxylase (ODC) has been shown to be induced by a number of conditions such as cold-injury, kindling, ischaemia and excitotoxin injection. In previous studies we have characterised the cortical response to kainate injection into the nucleus basalis and shown a substantial increase in both ODC mRNA and enzyme activity which reaches a maximum at 8h. This response is completely prevented by treatment with MK-801, indicating the involvement of NMDA receptors in mediating this response. Whilst NMDA receptors are known to gate a cation channel leading to increased calcium entry, an additional effect on the release of arachidonic acid has been reported. The possibility that NMDA receptor mediated activation of phospholipase A2 and release of arachidonic acid might mediate this ODC response was investigated in this study by treatment with the phospholipase inhibitors quinacrine and dexamethasone. Treatment of animals with quinacrine (100 mg/kg) at the time of injection of kainate into the nucleus basalis caused a significant attenuation of the induction of ODC in cerebral cortex of 43%. No further attenuation was seen at higher doses. A similar reduction in ODC induction was seen after treatment with dexamethasone (1 mg/kg) but a greater effect could be obtained (65% attenuation) at higher doses. The possible involvement of arachidonic acid derivatives in mediating ODC induction was further investigated by treatment with the cyclo-oxygenase inhibitor indomethacin and the lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA). Indomethacin was able to significantly attenuate the induction of ODC (greater than 60%) whilst NDGA (30 mg/kg) was ineffective. These results indicate the possible role of arachidonic acid derivatives in the regulation of the expression of ODC in cerebral cortex after excitotoxin injection.