Sensory, motor and cognitive alterations in aged cats.

These experiments were designed to assess some of the sensory, motor and cognitive alterations that occur in aged cats. Three groups of cats (1-3, 5-9 and 11-16 years of age) were tested in four behavioral tasks to assess age-dependent changes in locomotor activity, fine motor coordination, reactivity to auditory stimuli and spatial reversal learning. In tests of locomotor activity, 11-16 year old cats displayed altered patterns of habituation compared to 1-3 and 5-9 year cats. There were no decrements in fine motor coordination in the 11-16 year cats as measured by their ability to traverse planks of varying width or by their scores on a neurological examination. The 11-16 and 5-9 year cats both displayed increased reactivity to auditory stimuli. On tests of spatial reversal learning, 11-16 year cats displayed superior performance compared to 5-9 or 1-3 year animals, making fewer errors and requiring fewer trials to reach criterion. These findings indicate that a series of age-related behavioral changes occurs in the cat. Some of these may be related to morphological and neurophysiological alterations in neurons in the caudate nucleus.

Quantitative morphology of medium-sized caudate spiny neurons in aged cats.

These studies were designed to assess some of the morphological alterations that occur in medium-sized spiny neurons of the caudate nucleus in aged cats. Computer assistance was used to quantify in three dimensions the extent of the dendritic trees of 164 neurons from 11 cats (5 1-3 years and 6 over 10 years of age) stained by the rapid Golgi technique. In all animals beyond 10 years of age there was a decrease in the density of spines on distal dendritic segments. This decrease was moderate (16%) in 13 year old cats and reached about 50% in 15 and 18 year old animals. In addition, there was an increase in the frequency of occurrence of spines with enlarged heads in all aged cats. In cats over 13 years there was a marked loss of portions of distal dendritic segments. All measures of dendrite length displayed statistically significant decreases of 30-40% in cats 15 and 18 years of age. There were no significant age-related alterations in numbers of dendrites, number of branches per dendrite or soma diameter. These morphological results indicate that there is a sequence of age-related changes that occurs in caudate medium-sized spiny neurons and provides a basis from which to assess functional alterations.

Postnatal development of caudate input neurons in the cat.

Lectin-bound horseradish peroxidase (WG-HRP) was pressure-injected into the caudate nucleus (Cd) of neonatal (less than 24 hours of age) and adult cats in order to assess the postnatal development of monosynaptic Cd input neurons. Tissue was processed for peroxidase activity with a benzidine dihydrochloride chromagen. The injection of WG-HRP produced relatively similar labelled zones of marker uptake in the caudate nuclei of both neonates and adults. Similar axonal projections were also labelled in both age groups. While many characteristics of retrogradely labelled CD input neurons were apparently constant throughout postnatal life, each of these features had a particular developmental modification. (1) Regardless of age, neuronal somata that projected to the CD were located in the neocortex, thalamus, substantia nigra, mesencephalic raphe nuclei, and globus pallidus. In each of these brain sites, labelled CD input neurons appeared to migrate postnatally. (2) The Cd afferent axons originated from the same neuronal lines in neonates and adults--small-to-medium-sized cortical neurons and medium-sized-to-large fusiform cells in all other brain sites. In each of the brain sites, labelled neurons displayed marked postnatal somatic growth. (3) In both age groups, there was a characteristic intrasomatic reaction product density in the labelled neurons located in each brain site (substantia nigra greater than thalamus = raphe = globus pallidus greater than cortex). In each of these brain sites, the intrasomatic reaction product density was less in neonates than in adults.

GABAergic basal forebrain neurons project to the neocortex: the localization of glutamic acid decarboxylase and choline acetyltransferase in feline corticopetal neurons.

Our objective was to determine whether GABAergic and cholinergic basal forebrain neurons project to the neocortex. The retrograde connectivity marker wheat germ agglutinin lectin-bound horseradish peroxidase was injected into the neocortex of adult cats. Histo- and immunohistochemical methods were combined to label sequentially connectivity and transmitter markers (glutamic acid decarboxylase; choline acetyltransferase) in forebrain neurons. The labels of each marker were identified by correlative light and electron microscopy. Two principal types of doubly labeled neurons were demonstrated. The connectivity marker was colocalized with glutamic acid decarboxylase or choline acetyltransferase. The neurons were located in the basal forebrain. Their ultrastructural, cellular, and regional organization supported 2 conclusions. (1) GABAergic basal forebrain neurons project to the neocortex. This is important new morphological evidence for the origin of inhibitory neocortical afferents from a subcortical brain site. (2) The GABAergic and cholinergic basal forebrain neurons projecting to the neocortex exhibit remarkable structural similarities. The transmitter diversity of these intertwined neocortical afferents may be significant for the pathology and treatment of human neurological disorders such as Alzheimer's disease.

Patterns of tachykinin expression and localization in developing feline neostriatum.

The development of tachykinins in the neostriatum was determined qualitatively in order to characterize the ontogeny of an early-forming neostriatal peptidergic system. Tachykinins were detected by immunohistochemistry in fetal, postnatal, and adult cats. Neostriatal cells and neurites expressed tachykinins as early as fetal age 30 and increased in frequency progressively with age. Initial tachykinin expression occurred in neostriatal neurons during their postmitotic migration. In the head of the caudate nucleus, clusters of tachykinin-containing cells and fibers formed between fetal days 35 and 45, when the distribution of labeled neurons changed from a dispersed to an aggregated pattern. Between fetal days 45 and 50, tachykinin-rich neuronal clusters increased in frequency and were distributed throughout the rostral caudate nucleus. In contrast to neurons in clusters, neurons in the complementary neuropil expressed tachykinins largely postnatally. Postnatal morphological maturation of tachykinin-containing neurons paralleled the morphogenesis of medium spiny neostriatal cells. In addition, the caudate nucleus and putamen followed different spatiotemporal gradients of tachykinin expression. These results indicate that tachykinins are expressed in neostriatal neurons during the early ontogeny of the neostriatum and may function as trophic factors before synaptogenesis.

Neurophysiological, pharmacological and morphological properties of human caudate neurons recorded in vitro.

Tissue samples from the caudate nucleus were obtained from eight children (eight to 172 months of age) who underwent hemispherectomies for the relief of intractable seizures. Neurophysiological, pharmacological and morphological properties of caudate neurons were characterized by intracellular recordings in an in vitro slice preparation. These properties were compared with those of tissue obtained from animal studies. Electrophysiological properties of human caudate neurons that were similar to those of cat caudate and rat neostriatal cells included resting membrane potential, input resistance, action potential rise time, fall time, duration and action potential afterhyperpolarization amplitude, as well as the general characteristics of locally evoked synaptic responses. Properties that were different included action potential amplitudes and time-constants. Human caudate neurons also displayed responses similar to those of cat caudate or rat neostriatal cells to manipulation of excitatory amino acid receptor systems and to dopamine application. Kynurenic acid, a broad-spectrum excitatory amino acid receptor antagonist, decreased the amplitude of evoked synaptic responses, indicating that they were partially mediated by excitatory amino acids. In Mg2+ free Ringer's solution, the amplitudes and durations of postsynaptic responses were increased and bursts of action potentials were induced. These effects were mediated by activation of N-methyl-D-aspartate receptors since they were blocked by 2-amino-5-phosphonovalerate, a specific N-methyl-D-aspartate-receptor antagonist. Iontophoretic application of N-methyl-D-aspartate also induced membrane oscillations and bursts in almost all caudate neurons. Dopamine decreased the amplitude of postsynaptic responses, an effect antagonized by domperidone, a selective D2 dopamine receptor antagonist. Developmentally, the greatest change was an increase in action potential amplitude, although input resistance decreased and action potential afterhyperpolarization amplitude increased. Postsynaptic responses were similar across age. All but one of the caudate neurons identified by intracellular injection of biocytin or Lucifer Yellow were medium-sized spiny cells. These experiments show that human caudate neurons display a number of electrophysiological properties similar to rat neostriatal or cat caudate neurons recorded in brain slices. Furthermore, few electrophysiological parameters changed significantly over the age period examined suggesting that the human caudate at eight months displays many of the neuronal functions of the more mature caudate nucleus.

Age-induced changes in electrophysiological responses of neostriatal neurons recorded in vitro.

The present studies were undertaken to determine whether the major electrophysiological characteristics of neostriatal neurons are altered during aging. The passive and active membrane properties of 130 neostriatal neurons obtained from young (three to five months, N = 65) and aged (24-26 months, N = 65) Fischer 344 rats were compared using an in vitro slice preparation. The results indicated that in a population of aged neostriatal neurons the majority of the electrophysiological changes that occurred resulted in decreases in cellular excitability. These changes included increased threshold to induce action potentials by intracellular current injection and decreased negativity of membrane potentials at which such action potentials were induced. In addition, there were increases in the amplitude of the action potential afterhyperpolarization and increases in the frequency of occurrence of accommodation when trains of action potentials were induced. These two latter effects can limit the frequency of action potential generation. The thresholds to elicit synaptically evoked depolarizing responses and action potentials were increased. The results also indicated that a number of basic electrophysiological parameters were unchanged by the aging process. These included action potential amplitude, rise time and duration, resting membrane potential, input resistance and time constant. Although thresholds for the induction of synaptic and action potentials by extracellular stimulation were increased, the latency, amplitude and duration of the evoked depolarization remained unchanged. These findings suggest that the ability of neostriatal neurons to integrate spatiotemporal inputs must be severely compromised in this population of aged cells. Furthermore, the present findings, when compared with age-induced electrophysiological alterations in neurons in other brain areas, indicate that age may differentially alter electrophysiological properties of neurons in separate nuclei. Profiles of age-related changes in neurophysiological properties of neurons provide important information that can be related to the contributions of individual neural areas to the behavioral effects of aging.

Altered excitatory amino acid function and morphology of the cerebellum of the spastic Han-Wistar rat.

A mutant strain of Han-Wistar rat carries an autosomal recessive gene producing spastic paresis which is characterized by ataxia, tremor and hind limb rigidity. Brains of affected rats and unaffected littermate controls were transected at the mesencephalon into rostral and caudal portions (the caudal portion contained the cerebellum and brainstem). Poly(A)+ mRNA was isolated from pooled rostral or caudal portions and injected into Xenopus oocytes. The oocytes were voltage-clamped and exposed to 1 mM L-glutamate, 500 microM kainate, 500 microM quisqualate, 200 microM N-methyl-D-aspartate (NMDA) or 1 mM gamma-aminobutyric acid (GABA). Oocytes injected with mRNA isolated from the caudal portions of the affected rat brains exhibited statistically significant increases in glutamate and kainate peak current responses compared to oocytes injected with mRNA from other brain samples. No differences were noted in the responses of the groups when exposed to quisqualate, NMDA or GABA. Cerebellar and brain stem mRNA were also isolated separately in different groups of mutants and unaffected littermates. Only oocytes injected with cerebellar mRNA from mutants displayed statistically significant increases in responses to glutamate and kainate. In parallel morphological studies changes in the cerebellum of mutants were also observed. These consisted of a loss of Purkinje cells and an asymmetrical disarrangement of the granule cell layer of cerebellar cortex. Taken together, the physiological and morphological results suggest that alterations in glutamate/kainate receptors in the cerebellum are phenotypic manifestations of the Han-Wistar mutation. The results are consistent with the hypothesis that this mutant rat might serve as a model of glutamate/kainate excitotoxicity in the brain.

Intracellular studies of the convergence of sensory input on caudate neurons of cat.

Quantitative analyses of the intracellular responses to peripheral stimuli were made in a 172 neurons recorded in the head of the caudate nucleus (Cd) of cat. Responsiveness of Cd neurons was tested using auditory and somatosensory stimuli which were presented unilaterally. We found that most cells (99%) responded to both auditory and somatosensory stimuli. Excitation followed by inhibition (i.e. E-I response) was the primary pattern of intracellular response occurring approximately 75% of the time. Qualitative and quantitative analyses revealed marked similarities in the amplitudes and time courses of the intracellular responses following supramaximal stimulation of the peripheral stimuli studied. Similarly, analyses of 40 pairs of simultaneously recorded neurons revealed a marked homogeneity in the intracellular responses to the same stimulus even though the recordings were made several mm apart. The most striking feature of the simultaneous recordings was that the peaks of the initial E-I response and the afterpotentials often occurred in register after a peripheral stimulus. Furthermore, changes in the responses of pairs of simultaneously recorded neurons co-varied. Because the striatum typically demonstrates spontaneous rhythmic activity, these data suggest that a peripheral stimulus has the potential of briefly synchronizing Cd's neuronal activity.

Aging reduces somatosensory responsiveness of caudate neurons in the awake cat.

Neuronal activity was recorded in 7 awake cats (3 animals 11-15 years and 4 animals 1-3 years of age) to determine if the ability of caudate neurons to process facial somatosensory information was impaired in aged animals. Proportionately fewer neurons responded to somatosensory stimuli and facial receptive fields were larger in the aged cats. In addition, proportionately fewer caudate neurons responded to cortical activation, there were fewer excitatory responses and spontaneous firing decreasing in aged cats. These results provide additional evidence that excitability of caudate neurons is reduced in aged cats.

Postnatal development of identified medium-sized caudate spiny neurons in the cat.

The morphology of intracellularly recorded neurons in the cat caudate nucleus (Cd) was studied during postnatal development. After intracellular recording of evoked responses in these neurons, horseradish peroxidase (HRP) was injected iontophoretically through the recording micropipette. Fifty-eight Cd neurons in cats ranging from 6 days of age through adulthood were identified morphologically. All of the recovered Cd cells were medium-sized spiny neurons. The basic somatodendritic morphology of these neurons was evident in the youngest kittens. The most striking morphological change was the postnatal formation of an extensive local axonal collateral plexus. The development of these local axonal collaterals was also quantified with computer assistance in medium-sized Cd spiny neurons selected from silver-impregnated material. This analysis showed that the major development of the branches of this local plexus occurred between birth and 3-4 months of postnatal age. Data from both the HRP-filled and silver-stained axons indicated that the postnatal growth of the local axonal collaterals of the medium spiny cells was associated with the elaboration and increasing prevalence of evoked inhibitory postsynaptic potentials in Cd neurons.

Intracellular analysis of the development of responses of caudate neurons to stimulation of cortex, thalamus and substantia nigra in the kitten.

Intracellular recordings were made from caudate neurons in anesthetized kittens of 2-72 days of age. In adult cats, results of intracellular recordings indicate that caudate neurons respond most frequently to stimulation of their major afferents from cortex, thalamus and substantia nigra with a sequence of excitation followed by inhibition (EPSP-IPSP sequence). The results of the present study show that the prominent IPSP of this sequence is not well developed in young kittens and does not reach adult values in terms of frequency of occurrence until beyond 40 days of age. Amplitude and duration of the IPSP evoked by cortical stimulation also did not reach adult values until beyond 40 days of age. In contrast, EPSPs can be evoked in the youngest kittens by stimulation of afferents to the caudate. These findings suggest that the caudate nucleus may alter its role during development. In early postnatal periods it functions as a simple relay system transmitting incoming information to its outputs in a relatively unaltered fashion. Later in development it becomes a system capable of complex modulation and filtering of neural information.

The spontaneous firing patterns of forebrain neurons. IV. Effects of bilateral and unilateral frontal cortical ablations on firing of caudate, globus pallidus and thalamic neurons.

To assess the effects of partial deafferentation of the neostriatum on spontaneous neuronal activity in the basal ganglia and related thalamic nuclei, ablations of frontal cortex were carried out in adult cats. Postoperative measures of interspike intervals of single neurons in the caudate nucleus, globus pallidus and ventral anterior-ventral lateral complex of the thalamus revealed a slowing of neuronal firing in these structures as compared with non-lesioned controls. The fact that deafferentation by cortical damage produces changes in neuronal firing in target neurons of the striatum (globus pallidus) and in thalamic neurons at least two synapses removed from the striatum is noteworthy. The possible extent to which these results might have been influenced by reduction of cortical inputs to or denervation of the thalamus is discussed.

Projections to the neostriatum from the cat precruciate cortex. Anatomy and physiology.

The projections to the striatum from two cytoarchitectonically and functionally distinct subdivisions of the cat precruciate motor cortex were studied using anatomical and electrophysiological techniques. Our results indicate that the medial precruciate cortex (stimulation of which leads to movements of the axial and proximal musculature) has a widespread projection to the lateral half of the caudate nucleus. The lateral precruciate cortex (stimulation of which leads to movements of the distal musculature) has a localized projection within the caudate nucleus adjacent to the internal capsule. Both medial and lateral precruciate areas project to the putamen. These results are discussed in relation to recent studies suggesting that the basal ganglia are involved in the enabling and sequencing of movements.

Physiological and morphological analyses of ventral anterior and ventral lateral thalamic neurons in the cat.

Intracellular recordings were made from ventral anterior and ventral lateral (VA-VL) thalamic neurons in the cat. VA-VL neurons were tested for responsiveness to activation of cortical, pallidal and cerebellar afferents, and were identified morphologically by intracellular injection of HRP. Orthodromic activation of cortical and pallidal afferents produced primarily an initial inhibition (due in part to oligosynaptic circuitry) while activation of cerebellar afferents produced an initial excitation in the majority of neurons tested. Antidromic activation of thalamocortical relay neurons was observed in 32% of the neurons tested. Neurons showing short latency responses to activation of globus pallidus-entopeduncular nucleus and cerebellar peduncle were concentrated in the medical and ventral portions of the VA-VL complex, respectively. Neurons showing short latency responses to activation of the neocortex were located throughout the entire extent of the VA-VL complex. Only 3% of the neurons tested showed short-latency convergence of cortical, pallidal and cerebellar afferents. In contrast, 53% of neurons tested showed long latency triple convergence. Eight VA-VL neurons were stained intracellularly with HRP. Based on dendritic morphology, the labeled neurons were separated into two types: a stellate type with dendrites that spread radially from the soma, and a fusiform type with dendrites that were oriented mainly parallel to the long axis of the soma. Both types of neuron were aspiny although the dendrites of the stellate cells exhibited short appendages.

Effects of caudate nuclear or frontal cortical ablation in neonatal kittens or adult cats on the spontaneous firing of forebrain neurons.

In this paper we have determined the long-lasting consequences of caudate and frontal cortical lesions on spontaneous neuronal firing. Lesions were made both in neonatal and adult cats. All recordings were made in adults. Qualitatively, the effects of the caudate ablations were similar whether they had been carried out in kittens or in adult cats. Caudate lesions produced long-lasting (greater than or equal to 1 year) decreases in the spontaneous firing of cortical neurons. These changes were more pronounced when made in neonates than in adults. The distributions of mean interspike intervals were also altered by these caudate lesions in the pallidum and in the ventral lateral nucleus of the thalamus. Again these effects were more marked if the animals were lesioned as neonates than as adults. Frontal cortical lesions inflicted upon adult cats produced more widespread changes in spontaneous firing rates than similar lesions made in neonates. In both groups frontal lesions slowed spontaneous firing and changed the distributions of mean interspike intervals of caudate neurons. These effects were long-lasting (greater than or equal to 1 year in neonatally-ablated animals). Cortical lesions made in adult cats markedly altered thalamic and pallidal spontaneous activity. Similar lesions made in neonates produced relatively small changes in thalamic and pallidal activity.

Postnatal ontogeny of evoked neuronal responses in the substantia nigra of the cat.

Single unit extracellular responses evoked by striatal, cortical, and somatosensory stimulation were recorded in substantia nigra (SN) neurons of kittens (1-70 days of postnatal age) and adult cats. Neuronal responses to stimulation of each site were obtained throughout postnatal development. However, 4 major developmental shifts in the response of neurons were found: (1) the responsiveness of neurons to orthodromic activation increased with age; (2) the signs of the initial responses of neurons to orthodromic activation increased with age; (2) the signs of the initial responses of neurons to caudate and cortical stimulation changed with age (excitatory and inhibitory responses in kittens vs almost entirely inhibitory responses in adults); (3) the the response latencies of neurons decreased with age; (4) nigral neurons that were activated antidromically by stimulation of nigrostriatal axons showed age-related decreases in refractory periods and increases in conduction velocities.

Amphetamine alters evoked responses of nigral neurons in kittens and adult cats.

The effects of i.p. amphetamine administration (5 mg/kg) on the evoked unitary responses of substantia nigra (SN) neurons to electrical stimulation of their afferents were tested in 4 kittens (3-27 days of age) and 4 adult cats. In adults, amphetamine had two major effects: (1) it blocked temporarily (15-30 min) all neuronal responses caudate (Cd) and cortical (Cx) stimulation; neuronal responsiveness recovered by 75 min post-drug; and (2) after 15 min postdrug, Cd and Cx stimulation evoked initial excitatory responses that were almost never found predrug. The latencies of Cd-evoked excitations indicated the existence of a mono- or oligosynaptic excitatory strionigral pathway while latencies of Cx-evoked excitations suggested that corticonigral excitatory influences were mediated multisynaptically. In kittens, amphetamine also produced an initial blockade of Cd- and Cx-evoked responses. However, the sign of initial responses to Cd stimulation was not altered since excitations were found both before and after drug treatment. These results indicated that amphetamine reveals excitatory evoked responses of SN neurons to striatal and striatally-mediated inputs that are masked during the course of normal postnatal development. Drug-related alterations of afferent inputs to SN neurons may underlie amphetamine-induced shifts in spontaneous neuronal activity which have been reported frequently.

The GABAergic striatonigral neurons of the cat: demonstration by double peroxidase labeling.

GABAergic striatonigral neurons were demonstrated in the adult cat by the specific double peroxidase labeling of a transmitter marker with an agranular appearance (GAD, the synthetic enzyme of GABA) and a connectivity marker with a granular appearance (WGA-HRP). Each marker was associated with different organelles confined to the perikaryal cytoplasm of neurons. GABAergic striatonigral neurons were of medium size, high frequency and wide location in the rostral caudate nucleus and putamen based on correlative light and electron microscopic identification. These cells had somatic and/or proximal dendritic spines and folded nuclear envelopes in some cases. They received GABAergic axosomatic and axodendritic inputs with symmetric synaptic specializations. They were also contacted by axosomatic, axodendritic and axospinous terminals with asymmetric synaptic specializations. These results indicate that the GABAergic striatonigral neurons are, for the most part, medium spiny cells that also emit intrastriatal axonal collaterals. Their intra- and extrastriatal axons mediate inhibitory postsynaptic influences on their targets. Their degeneration might contribute to the GABAergic deficits found in the basal ganglia in Huntington's disease.

Branched projections of pallidal and peripallidal neurons to neocortex and neostriatum: a double-labeling study in the cat.

Double-labeling of basal forebrain neurons by retrograde axonal transport of different markers demonstrated afferents shared by the neocortex and neostriatum. A considerable double-labeled complement of neurons located in the globus pallidus (lateral pallidal segment) and the adjacent interdigitating basal nucleus of Meynert (peripallidal region) had branched axonal collaterals projecting to the precruciate, cingulate and prorean gyri as well as to the head of the caudate nucleus.