C. Shaw

Nicotine receptors are located on lateral geniculate nucleus terminals in cat visual cortex

Using the methods of in vitro receptor autoradiography, we have characterized a population of receptors for nicotine in cat visual cortex that is concentrated primarily in layer IV of areas 17 and 18. Surgically undercutting the visual cortex essentially abolished [3H]nicotine binding in the isolated zone. However, neuron-specific, quinolinic acid lesions of a region of visual cortex had little effect on binding, establishing a presynaptic locus on cortical inputs for these sites. Lesions of the lateral geniculate nucleus abolished binding in the corresponding cortical areas, thus localizing the [3H]nicotine binding sites to lateral geniculate nucleus terminals in the cortex.

Ontogenesis of muscarinic acetylcholine binding sites in cat visual cortex: reversal of specific laminar distribution during the critical period☆

Acetylcholine appears to act as a modulator of neuronal activity in cat visual cortex and, like noradrenaline, may be involved with cortical plasticity mechanisms during the critical period. To explore possible ACh involvement in these mechanisms we have examined acetylcholine binding sites in cat visual cortex during development using [3H]QNB, a muscarinic antagonist. At 3 days postnatal [3H]QNB preferentially labelled binding sites in layer IV. During development the pattern of binding reversed, so that by 95 days postnatal layer IV was the least densely labelled. The number of binding sites increased during development peaking at 1 month postnatal. The Kd of [3H]QNB binding sites increased to 95 days postnatal, with a peak value of 0.76 nM. The results show that during development, and especially within the critical period, changes in [3H]QNB binding site distribution, number and affinity occur.

Ontogenesis of muscimol binding sites in cat visual cortex

In vitro receptor binding techniques were used to study the characteristics, distribution, and ontogenesis of muscimol binding sites in cat visual cortex. [3H] Muscimol, a GABA agonist, labelled a single population of binding sites with a KD of 18 nM in adult cats. Specific binding was saturable, reversible and was blocked by the addition of GABA or (+) bicuculline. Autoradiograms revealed that the highest density of [3H] muscimol binding occurred in cortical layer IV. Similar patterns of [3H] muscimol binding were observed at all ages examined, although the binding densities differed. The peak [3H] muscimol binding density, corrected for amount of protein, occurred at 3 months postnatally. In 3 day old and adult cats binding density was 41% and 69%, respectively, of the peak value.

Characteristics and distribution of muscimol binding sites in cat visual cortex.

In vitro receptor binding techniques were used to study the characteristics and distribution of [3H]muscimol binding sites in cat visual cortex. [3H]muscimol, a specific GABA agonist, labeled a single population of binding sites with a Kd of 18 nM. Specific binding was saturable, reversible, and was blocked by the addition of GABA or (+)-bicuculline. Autoradiograms revealed that the highest density of [3H]muscimol binding sites occurred in cortical layer IV. Little variation between the various visual cortical areas was noted in contrast to marked regional heterogeneity within subcortical structures.

The neural circuitry of the neocortex examined in the in vitro brain slice preparation

The in vitro brain slice technique has been applied to the study of the neocortex. Cortical blocks were removed from adult rats deeply anesthesized with halothane, sectioned coronally at 400-700 micrometer, and placed in a brain slice chamber. Cortical slices typically showed spontaneous and evoked potential activity and normal histology for 8 h or longer. Single units and evoked potential recordings were made from different layers of the cortex using micropipettes. The evoked potentials to electrical stimulation of differing intensity, frequency, and from different cortical layers were analyzed. Evoked potential from all but the most superficial layers of the cortex showed a characteristic 6-component response to stimulation of nearby white matter. This evoked potential closely resembled cortical responses recorded in vivo by other investigators following afferent stimulation. The response amplitude of all components increased as stimulus intensity was raised. Radial movement of the recording electrode showed that components 1-3 had their largest amplitudes in the deepest cortical layers, component 4 reached its greatest amplitude and shortest latency in layer IV, and components 5 and 6 reached their greatest amplitudes in layers IV to II. The frequency following for various components was measured showing greater decline in amplitude for components 4-6 than 1-3. This, together with the results of previous investigators, suggests that the first 3 components represent afferent fiber input, while component 4 represents the first cortical response (layer IV). Components 5 and 6 represent later, additional cortical responses. Further support for the intracortical origin of component 4 was provided by lateral intracortical stimulation within layer IV, giving an evoked potential composed mostly of component 4. With lateral movement of the recording electrode in layer IV the evoked potential disappeared in under 1 mm, suggesting a fairly restricted afferent input to the cortex. The present results encourage the use of the cortical brain slice preparation as an appropriate model system in which to study cortical neural circuitry.

Ontogenesis of β-adrenergic binding sites in kitten visual cortex and the effects of visual deprivation

We have determined maximum binding capacities for beta-adrenergic binding sites in developing cat visual cortex. These increase quickly from birth to 4 weeks, after which time a slower increase reaches maximal levels at 12 weeks. Dark-rearing and monocular eyelid suture have no influence on this developmental profile.

Modification of neurotransmitter receptor sensitivity in cat visual cortex during the critical period

We have examined the characteristics of various receptors in cat visual cortex during postnatal development. These included beta-adrenergic, GABA, benzodiazepine and acetylcholine receptors. For each population of receptor the number (Bmax) and affinity (Kd) were examined as a function of postnatal age (3 days-adult). For all receptors examined, the Bmax increased during development from low early values to a peak within the critical period. The Kd also changed during development for most receptors. The simultaneous alterations in Bmax and Kd necessitate defining a term which takes both of these receptor properties into consideration. This term, called receptor sensitivity (RS), provides a more comprehensive measure of receptor function than either Bmax or Kd alone. Using this measure, we find that receptor sensitivity is low near birth for the 4 receptor populations studied, rises to a peak within the first two months of life, and then declines to near-neonatal levels for 3 of the 4 receptor populations.

The distribution and ontogenesis of [3H]nicotine binding sites in cat visual cortex.

In vitro autoradiographic techniques using [3H]nicotine were used to characterise nicotine binding sites in developing kitten visual cortex. These binding sites in adult animals have a Bmax of 3.91 fmol/mg protein and a Kd of 4.40 nM. Displacement experiments indicate that [3H]nicotine binds to a nicotinic receptor site that is similar to central nicotinic sites described by investigators in other mammals. The number of binding sites increases during postnatal development, peaking near 60 days of age and levelling-off thereafter. There is no evidence for large changes in affinity during postnatal development for this binding site. [3H]Nicotine binding sites are densely concentrated in layer IV in the visual cortex of adult animals, with sharply reduced binding outside of cortical areas 17 and 18. This laminar pattern does not change during postnatal development, but an increase in the number of binding sites in layer IV as well as in layers I and VI occurs during early postnatal life. These binding sites disappear when extrinsic cortical inputs are severed. However, they survive when neurons in the visual cortex are selectively destroyed with a cell-specific neurotoxin. Unilateral destruction of the lateral geniculate nucleus eliminates [3H]nicotine binding sites in the visual cortex ipsilateral to the lesion, suggesting that they are located presynaptically on the terminals of lateral geniculate nucleus afferent fibres. The laminar pattern of binding of [3H]nicotine during early development of the visual cortex is complimentary to that for muscarinic acetylcholine receptors. These latter receptors redistribute during postnatal development becoming less prominent in layer IV at the same time as the [3H]nicotine binding sites are increasing in number in this layer. For a short period of time at the height of the critical period for cortical plasticity, both populations of binding sites are located in layer IV.

Alterations in receptor number, affinity and laminar distribution in cat visual cortex during the critical period.

The number, affinity, and laminar distributions of various receptors in cat visual cortex were examined during postnatal development using homogenate and in vitro autoradiographic techniques. For all receptor populations examined, the total number of receptors (Bmax) increased from relatively low early values to peak values during the first three months of postnatal life followed by a drop or plateau in the number of receptors. This peak in Bmax occurred during the physiologically-defined period for cortical plasticity. For most receptors examined, the affinity (KD) was also altered during postnatal development. Many of the receptor populations examined exhibited changes in their initial laminar distributions during the first three months of postnatal development, although other did not. The results show a more complex picture of receptor ontogenesis than previously reported, and suggest that the observed receptor modifications affect the synaptic efficacy and the basic chemical circuitry of the visual cortex during the critical period.

Development of phorbol ester (protein kinase C) binding sites in cat visual cortex

Tritiated phorbol-12,13-dibutyrate [( 3H]PDBu), a phorbol ester, was utilized to autoradiographically localize protein kinase C (PKC) in the cat visual cortex. Thin, slide-mounted sections of adult cat brain were used to characterize binding of [3H]PDBu. This was found to be saturable, reversible, and more readily displaced by phorbol ester than by synthetic diacylglycerols. Binding sites displayed a tissue concentration of 20 pmol/mg protein, and a dissociation constant of 8.0 nM. [3H]PDBu was slow to associate with its receptor, requiring 9.5 h to reach equilibrium. Autoradiograph revealed that PKC is heterogeneously distributed in the cat brain, and displays a laminar-specific pattern in the visual cortex. This laminar distribution undergoes marked changes during the first two months of postnatal life. In the visual cortex of neonatal kittens, [3H]PDBu binding is confined to layers I and V. Layer III acquires high levels of binding by postnatal day 15, layer II by 28 days, and layer VI becomes labelled by 40 days of age. Adult animals exhibit high levels of binding in all laminae except layer IV. Age-dependent changes in PKCs laminar distribution do not seem to be correlated with specific anatomical, neurochemical, or behavioural events during development. PKC appears to be associated with cell bodies or processes intrinsic to the visual cortex, and is probably not located on the terminals of cortical afferents.