John S. McKenzie

Localization of binding sites for calcitonin gene-related peptide in rat brain by in vitro autoradiography

The distribution of binding sites for calcitonin gene-related peptide (CGRP) in rat brain were studied using in vitro autoradiography. In a radioreceptor assay using [125I]human calcitonin gene-related peptide as the radioligand, with cerebellar cortical membranes, rat calcitonin gene-related peptide had a binding affinity constant of 1.16 +/- 0.23 X 10(10) M-1 and a site concentration of 43.4 +/- 3.4 fmol/mg protein. In this system, human calcitonin gene-related peptide had a binding affinity constant of 3.9 +/- 0.7 X 10(9) M-1 whereas salmon calcitonin was very weak with a binding affinity constant of only 6.8 +/- 4.0 X 10(5) M-1. CGRP binding localized by in vitro autoradiography, using [125I]rat calcitonin gene-related peptide, had a characteristic distinct distribution in the rat brain. There were high concentrations of binding found over the accumbens nucleus, the organum vasculosum of the lamina terminalis, ventral caudate putamen, median eminence, the arcuate nucleus, lateral amygdaloid nucleus and lateral mammillary nucleus, the superior and inferior colliculi, pontine nuclei, molecular and Purkinje cell layers of the cerebellar cortex, the nucleus of the solitary tract, the inferior olivary nuclei, hypoglossal complex and the vestibular and cochlear nuclei. The distribution of these binding sites suggests multiple roles for CGRP in the central nervous system including auditory, visual, gustatory and somatosensory processing, and in neuroendocrine control.

Evidence for a new subclass of calcitonin/calcitonin gene-related peptide binding site in rat brain

Binding sites for calcitonin and calcitonin gene-related peptide are widely distributed in the central nervous system. In this study, binding of [(125)I]-alpha-rat calcitonin gene-related peptide and [(125)I]-salmon calcitonin in adjacent sections of rat brain revealed clearly distinct patterns of binding in most regions although in some restricted areas such as parts of the ventral striatum, including the nucleus accumbens, there was some overlap in the patterns of binding. In the primary olfactory cortex, which bound only calcitonin gene-related peptide, salmon calcitonin was very weak in inhibiting the binding of calcitonin gene-related peptide. In the nucleus accumbens, high affinity binding of calcitonin and calcitonin gene-related peptide at their homologous receptors was observed, with affinity constants for calcitonin and calcitonin gene-related peptide of 1.4 x 10(9) M(?1) and 1.2 x 10(9) M(?1) respectively. Cross competition studies in this nucleus demonstrated that salmon calcitonin was able to compete for [(125)I]-rat calcitonin gene-related peptide labelled sites with high affinity, with an affinity constant of 0.8 x 10(9) M(?1). However, rat calcitonin gene-related peptide was less potent in inhibiting the binding of [(125)I]-salmon calcitonin labelled sites with only 28% inhibition at 10(?6)M. Further characterization of the calcitonin sensitive calcitonin gene-related peptide labelled sites demonstrated that a range of calcitonin analogs inhibited the binding of [(125)I]-rat calcitonin gene-related peptide with the same order of potency as the analogs competed for [(125)I]-salmon calcitonin labelled sites. Digital substraction mapping revealed calcitonin-sensitive calcitonin gene-related peptide binding sites over parts of the ventral striatum, including mid-caudal nucleus accumbens and fundus striati; over the lateral border of the lateral bed nucleus of the stria terminalis; part of the central amygdaloid nucleus; the organum vasculosum of the lamina terminalis and area postrema and over the wings of the dorsal raphe. These results demonstrate the existence of a new subtype of calcitonin/calcitonin gene-related peptide binding site, which has high affinity for the two otherwise biochemically distinct peptides.

Effects of lesions in the horizontal diagonal band nucleus on olfactory habituation in the rat

The nucleus of the horizontal limb of the diagonal band, a component of the basal forebrain magnocellular complex affected in Alzheimer type dementia, supplies centrifugal innervation to the olfactory bulb. We have tested the hypothesis that horizontal limb of the diagonal band lesions will interfere with olfactory memory in a simple olfactory test paradigm. Lesions occupied a restricted volume, approximately 20%, of medial horizontal limb of the diagonal band. There was interference with habituation of investigation latency and duration, six and 16 days after lesioning. It is concluded that bilateral partial lesions of the medial nucleus of the horizontal limb of the diagonal band interfere with habituation memory for odours.

Intracellular responses of olfactory bulb granule cells to stimulating the horizontal diagonal band nucleus

The effects of centrifugal afferents on membrane potentials of identified granule cell layer using evoked field potential profiles, and trans-synaptic activation via antidromic stimulation of output cell axon collaterals. Intracellular recordings maintained for 4-30 min showed complex spontaneous spike discharges and allowed characterization of the cells input resistance, and on some occasions its morphology following intracellular injection of Lucifer Yellow. Stimulation in the nucleus of the horizontal limb of the diagonal band, but not surrounding regions, produced hyperpolarizing responses in 13 of 27 cells in the granule cell layer; four of these were morphologically identified as granule cells of two types, in five the responses had reversal potentials more negative than the resting potential, and six were identified as granule cells by monosynaptic activation from output axon collaterals. A different set of three cells in the granule cell layer responded with depolarization. The results are consistent with the inhibition of tonic activity of granule cells by the nucleus of the horizontal limb of the diagonal band, leading to disinhibition of mitral and tufted cells via dendrodendritic synapses of granule cells on mitral/tufted cell secondary dendrites.

Effect of stimulating the nucleus of the horizontal limb of the diagonal band on single unit activity in the olfactory bulb

The effects of centrifugal afferents on single unit discharge in the main olfactory bulb were studied in anaesthetized rats. Recording with extracellular micropipettes revealed spontaneous firing in all bulb layers. Units were located to different laminae using evoked field-potential profiles and histological verification. Output neurons were identified by antidromic response to stimulation of the lateral olfactory tract. Single- or brief multiple-pulse stimulation in the nucleus of the horizontal limb of the diagonal band, but not in adjacent regions, facilitated 17 out of 27 mitral cells with no effect on 10, but inhibited 21 out of 33 granule cell layer units with no effect on 12. Of 13 presumed tufted cells, six were facilitated and the rest unaffected. In contrast, stimulation of olfactory cortex inhibited mitral cells and facilitated most granule layer cells. The results are consistent with an inhibition of tonic granule cell discharge by the horizontal diagonal band nucleus, with resultant disinhibition of mitral cells via the dendrodendritic synapses of granule cells on mitral cell secondary dendrites.

Catecholamine uptake sites in mouse brain: distribution determined by quantitative [3H]mazindol autoradiography

Because of the importance of the mouse brain catecholamine system in the study of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and because little information is available concerning the chemical neuroanatomy of the mouse, catecholamine uptake sites were mapped in C57 black mouse brain using [3H]mazindol autoradiography. Displacement studies with known dopamine (DA) and noradrenaline (NA) uptake blockers showed that binding in the striatum was entirely to DA uptake sites, while binding in the locus coeruleus was to NA uptake sites only. By using the selective noradrenergic uptake blocker desmethylimipramine (DMI), a complete map of both DA and NA uptake sites was generated. The mesostriatal DA system was the most clearly labelled and uptake sites were seen better in striatal terminals than the substantia nigra. Within the noradrenergic system, highest binding levels were seen over the locus coeruleus, although it was unclear whether these uptake sites were on cell bodies or terminals from the lateral tegmental noradrenergic system. These maps of the catecholamine uptake system in mouse brain provide a baseline for study of newly discovered neurotoxins and ageing processes.

Hippocampal suppression of intralaminar thalamic unit responses in cats and comparison with suppression evoked from the caudate nucleus

Abstract Conditioning electrical stimuli applied to the dorsal hippocampus of chloralose-anaesthetized, paralyzed cats, suppressed somatically evoked unit responses in the intralaminar thalamus. Mean onset latency for suppression was 29 msec (range: 10–45 msec), and mean duration of complete response suppression was 318 msec (range: 125–700 msec). Electrical stimulation at the level of the hippocampal pyramids or their proximal dendrites was required for suppression of intralaminar responses. Stimulation of the dentate fascia and area CA4 did not exert the same effect. The effect was not due to spread of current to supraventricular tissue, and was blocked by interruption of the fornix outflow pathway. Unit responses were suppressed by hippocampal stimulation more frequently in rostral centrolateral than in caudal centromedial and parafascicular intralaminar areas. A proportion of units in adjacent areas of laterodorsal, dorsomedial, ventrolateral, lateral posterior and limitants nuclei, and in the prerubral field, were also suppressed. No ventrobasal units, lemniscal or extralemniscal, were suppressed. Many of the same intralaminar units were suppressed by either hippocampal or caudate nucleus stimulation. Caudate stimulation was more likely than hippocampal to suppress units in caudal intralaminar nuclei. Suppression of intralaminar somatosensory responses is considered to provide a possible neural basis for reported similarities in the involvement of the hippocampus and caudate nucleus in behavioral plasticity.

An electrophysiological study of thalamo-caudate neurones in the cat.

SummaryThalamo-caudate projection neurones were identified in cats by antidromic activation from the caudate nucleus, under chloralose anaesthesia or in cerveau isolé preparations.Units in nuclei centralis lateralis, medialis dorsalis, and centrum medianum-parafascicularis responded antidromically to one or more electrodes of a caudate nucleus array at latencies ranging from 0.4 to 16 ms. The responses did not appear to result from stimulus spread to the internal capsule.A separate population of medial thalamic units responded post-synaptically to caudate stimulation, at modal latency exceeding modal antidromic latency by 2 ms. Comparison of apparent conduction velocities for each thalamic nucleus indicated that the postsynaptic responses were activated by collaterals of thalamo-caudate neurones.A proportion of thalamo-caudate neurones responded to somatic stimulation; some of these were inhibited by caudate stimulation, and a few discharged on substantia nigra stimulation. These results demonstrate the possibility of somatic afferent and nigrofugal inputs to the caudate nucleus mediated by neurones of the medial thalamus.

Olfactory bulb output neurons excited from a basal forebrain magnocellular nucleus

We present intracellular data which demonstrates a unique facilitatory centrifugal influence on the output cells of the olfactory bulb; the source being the lateral component of the nucleus of the horizontal limb of the diagonal band (HDB), part of the basal forebrain magnocellular complex. Damage to this facilitatory HDB influence may explain the loss of olfactory sensitivity seen early in Alzheimers disease in which pathological changes occur in the basal forebrain.

Lesions in the magnocellular preoptic nucleus decrease olfactory investigation in rats

The nuclear complex of the horizontal limb of the diagonal band and the magnocellular preoptic nucleus, components of the basal forebrain magnocellular system affected in Alzheimer-type dementia, supply centrifugal innervation to the olfactory bulb. The lateral magnocellular preoptic nucleus provides significant GABAergic input. Since its stimulation may facilitate olfactory bulb mitral cells, we have investigated the effect of sub-total electrolytic lesions in this nucleus on performance in a simple test of olfactory investigation and its habituation. Two groups of rats used with lesions which occupied restricted volumes, approximately 30 and 15% of the magnocellular preoptic nucleus. Behaviorally, there was interference with olfactory investigation, with increased investigation latency and decreased investigation times, the group with larger lesions at 6 and 16 days after operation. There was no significant effect of the smaller lesions. No effects on patterns of olfactory habituation or discrimination were seen. The impairment of olfactory investigation could not be explained by interruption of medial forebrain bundle fibres traversing the nucleus. It is suggested that bilateral partial destruction of magnocellular preoptic neurones may produce significant deficits in either olfactory sensitivity or olfactory motivation.