Enhancement of conditioned arterial pressure responses in cats after brainstem lesions.

Cats were classically conditioned after the baroreceptor reflexes were abolished by bilateral placement of electrolytic lesions in the nucleus tractus solitarii. The conditioned increases in arterial pressure were more than five times larger than the responses obtained in similarly trained controls. This finding suggests that the baroreceptor reflexes actively inhibit conditioned increases of arterial pressure.

Local cerebral blood flow increases during auditory and emotional processing in the conscious rat.

Local cerebral blood flow was measured in rats by the 14C-labeled iodoantipyrine technique with quantitative autoradiography during the processing of environmental stimuli. Presentation of a tone increased blood flow in the auditory but not the visual pathway. When the animal had previously been conditioned to fear the tone, blood flow additionally increased in the hypothalamus and amygdala. Local cerebral blood flow can thus be used to detect patterns of cerebral excitation associated with transient (30- to 40-second) mental events in experimental animals.

Strain differences in pituitary prolactin content: relationship to number of hypothalamic dopamine neurons.

BALB/cJ mice have more tuberoinfundibular dopamine neurons, and thus greater tyrosine hydroxylase activity, than CBA/J mice. Strain differences in the synthesis and release of prolactin would also be predicted since dopamine released from the tuberoinfundibular neurons is the prolactin inhibitory factor which plays a role in the regulation of both prolactin synthesis and release. As expected, CBA/J mice, with fewer dopamine neurons, synthesized and released significantly more prolactin than BALB/cJ mice; that is, both pituitary and serum prolactin concentrations were greater in CBA/J mice. To determine if there were more cells containing prolactin or more prolactin per cell, pituitaries were stained with antibodies to prolactin and densitometric analysis made of both the average staining per unit area and total staining per pituitary. For both indices CBA/J mice had more staining than BALB/cJ mice. Using these criteria the difference in staining was attributed to more prolactin-stained lactotrophs in the CBA/J strain. Although no differences in the number of acidophils demonstrated by Pearse Trichrome method were observed, acidophils from BALB/cJ mice appeared smaller and contained less cytoplasm than those from CBA/J mice. We conclude that strain differences in the number of tuberoinfundibular dopamine neurons are inversely related to the number of immunocytochemically demonstrable prolactin-containing cells in the anterior pituitary.

Quantitative immunocytochemistry of tyrosine hydroxylase in rat brain. I. Development of a computer assisted method using the peroxidase-antiperoxidase technique.

We sought to develop a quantitative immunocytochemical procedure using the peroxidase-antiperoxidase (PAP) technique to analyze amounts of neurotransmitter biosynthetic enzyme tyrosine hydroxylase (TH) within the nucleus locus coeruleus (LC) of rat brain. Rats were perfused with 4% paraformaldehyde. The brains were embedded in paraffin, sectioned at 5 micrometers in the sagittal plane and immunocytochemically stained with antibodies to bovine adrenal TH. Staining intensity, measured by a TV image analysis system was reproducible within +/- 5%. Reaction conditions required so that the intensity of the PAP reaction product was directly and linearly related to the amount of TH enzyme protein in tissue were obtained by reacting tissues with saturating concentrations (2.5 mM) of diaminobenzidine (DAB) substrate, a constant dilution of antibody, and incubation time adjusted so that the darkest elements in tissue were below saturation. Variations in staining intensity of serial sections through the LC were found to be insignificant when compared to variations in staining between animals. In order to increase the amount of immunoreactive TH in the LC, groups of rats were treated with reserpine. Immunocytochemical and biochemical analyses were performed in parallel groups of control and reserpine animals allowed to survive 1-3 days following a single injection (10 mg/kg s.c.) of reserpine. A close correlation was found to exist between the amount of TH enzyme protein determined biochemically and the density of staining for TH. The maximal increase in TH measured immunocytochemically was 2.2-fold which was about 80% of the maximal induction determined biochemically. We conclude that the PAP method can be used for quantitative immunocytochemistry of brain TH providing that optimal reaction conditions are established.

Quantitative immunocytochemistry of tyrosine hydroxylase in rat brain. II. Variations in the amount of tyrosine hydroxylase among individual neurons of the locus coeruleus in relationship to neuronal morphology and topography.

We sought to determine by a computer assisted quantitative immunocytochemical method using the peroxidase-antiperoxidase (PAP) technique if individual neurons of the nucleus locus coeruleus (LC) varied with respect to the amount of tyrosine hydroxylase (TH) within each cell and, if so, whether the amount of TH is related to morphometric characteristics and/or location of the neuron within the LC. Appropriate immunohistochemical reaction conditions were used so that the intensity of DAB reaction product formed in the PAP reaction was directly and linearly related to the amount of TH enzyme protein present in the tissue. The distribution of the average optical density of 1425 individual LC neurons obtained from 5 animals and standardized to an overall mean density varies unimodally over a 3-fold range. The relationship between staining intensity and morphological and topographic characteristics of individual neurons was examined. There was no correlation between staining intensity and morphological features of individual neurons, including cytoplasmic area. However, differences in the amount of TH in individual neurons correlated significantly (P less than 0.01) with the location of the cell in LC: the lightest staining neurons were located in the central core of the LC. The density of staining did not differ significantly in the other 4 traditionally defined regions of LC (the anterior, posterior, and ventral poles of the LC and the nucleus subcoeruleus) which form a rim around the central core. Similarly, cells which are closest to the center of mass of the LC showed a lighter staining intensity than cells located progressively further from the center. These lighter staining cells in the center were also packed more closely together (P less than 0.01). Therefore dense packing of neurons correlates with light staining intensity (P less than 0.01). We conclude that neurons of the LC vary with respect to the amount of TH, which in turn relates to the location of the cell in the LC and possibly its packing density, but not its morphometric characteristics. Since in noradrenergic neurons, the amount of TH is directly influenced by firing rate, the results suggest that the biochemical heterogeneity of LC neurons reflects differences in their functional activities. In turn, this may relate to heterogeneity of LC with respect to projection fields.

Global increase in cerebral metabolism and blood flow produced by focal electrical stimulation of dorsal medullary reticular formation in rat.

We sought to determine whether the increases in local cerebral blood flow (LCBF) elicited by focal electrical stimulation within the dorsal medullary reticular formation (DMRF), are secondary to or independent of, increased local cerebral glucose utilization (LCGU). Rats were anesthetized (chloralose), paralyzed, artificially ventilated and arterial pressure and blood gases controlled. LCBF and LCGU were determined in two separate groups of animals, using the autoradiographic [14C]iodoantipyrine and [14C]2-deoxyglucose methods, respectively. In unstimulated controls, LCBF (n = 5) and LCGU (n = 5) were linearly related (r = 0.780; P less than 0.001) in the 27 brain regions studied. During DMRF stimulation LCGU increased significantly in 21 of the 27 regions, including cerebral cortex (up to 168% of control), thalamic nuclei (up to 161%) and selected ponto-medullary regions (e.g. parabrachial complex: 212%; vestibular complex: 147%). Along with LCGU, LCBF rose significantly in 25 regions (sensory motor cortex: 163%; anterior thalamus: 161%; parabrachial complex: 186%). Correlation analysis demonstrated that, during DMRF stimulation, the close relationship between LCBF and LCGU is preserved (r = 0.845; P less than 0.001) and that, in addition, the increase in LCBF (delta LCBF) is proportional to the increase in LCGU (delta LCGU) (delta LCBF = 2.18 delta LCGU + 6.92; r = 0.7729; P less than 0.001). Excitation of neurons or fibers within DMRF increases brain metabolism globally and blood flow secondarily. The DMRF appears to modulate cerebral metabolism globally, by as yet undefined pathways.

Electrical stimulation of cerebellar fastigial nucleus increases cerebral cortical blood flow without change in local metabolism: evidence for an intrinsic system in brain for primary vasodilation.

We sought to determine whether the increase in regional cerebral blood flow (rCBF) elicited by electrical stimulation of the fastigial nucleus of the cerebellum (FN) is secondary to, or independent of, increased local cerebral metabolism (rGMR) in anesthetized (chloralose) paralyzed rats. rCBF and rGMR were determined autoradiographically in separate groups of animals with comparable blood gases and systemic pressure, by the [14C]iodoantipyrine and [14C]2-deoxyglucose methods respectively. In sham-operated controls, rCBF (n = 5) and rGMR (n = 5) were closely related in the 28 brain areas studied (r = 0.733; P less than 0.005). During FN stimulation, rCBF (n = 6) increased significantly in 24 of the 28 areas, the greatest increase being in the cerebral cortex (up to 215%). rGMR (n = 9) increased in only 15 areas, so that the correlation between rCBF and rGMR throughout the brain became weaker (r = 0.568; P less than 0.005). Where the rCBF increases were the greatest (particularly in the cerebral cortex), rGMR was unchanged. This suggests that the rCBF increase was not a consequence of the increased rGMR. We conclude that neurons originating in or passing through FN may influence local cerebral circulation through a primary cerebral vasodilatation not coupled to metabolism.

Arterial pressure and heart rate changes during natural sleep in rat.

Mean arterial pressure and heart rate data during quiet wakefulness and phases of sleep in conscious rat are sampled by a computer at a rate of 100/sec. Average values and variability expressed as standard deviation are computed for each recording session. Mean arterial pressure and heart rate and their variability decrease from quiet wakefulness to synchronized sleep. During desynchronized sleep, mean arterial pressure increases to the level of quiet wakefulness, and is more variable than during synchronized sleep. Heart rate is lower and more uniform during sleep than during quiet wakefulness, and there is no difference between synchronized and desynchronized sleep except that a greater variability occurs during desynchronized sleep. The study shows that characteristic and specific cardiovascular changes accompany the phases of sleep and that a hierarchy of arterial pressure is present during the resting behavior in rat.

Binary tree representation of three-dimensional, reconstructed neuronal trees: a simple, efficient algorithm.

A simple, efficient algorithm is presented for the transformation of dendritic trees to binary tree representations. The binary tree form significantly reduces traversal time, simplifies software for tree analysis and yields compact storage of the neuronal trees. These algorithms are easy to program and are useful in quantitative neuroanatomical studies.

Role of local neurons in cerebrocortical vasodilation elicited from cerebellum.

The vasodilation elicited in cerebral cortex by stimulation of the cerebellar fastigial nucleus (FN) is mediated by input pathways coming from the basal forebrain. We studied whether these pathways mediate the cortical vasodilation via a direct action on local blood vessels or via interposed local neurons. Neurons were destroyed in the primary sensory cortex by local microinjection of the excitotoxin ibotenic acid (IBO) (10 micrograms/l microliter). Five days later rats were anesthetized (alpha-chloralose), paralyzed, and ventilated. Arterial pressure and blood gases were controlled, and FN was stimulated electrically. Local cerebral blood flow (LCBF) was measured using the [14C]iodoantipyrine technique with autoradiography. Five days after IBO, neurons were destroyed in a restricted cortical area, and afferent fibers and terminals were preserved. The selectivity of the neuronal loss was established by histological and biochemical criteria and by transport of horseradish peroxidase from or into the lesion. Within the lesion, resting LCBF (n = 7) was unaffected, but the increase in LCBF evoked from the FN was abolished (P greater than 0.05); n = 6). In contrast the vasodilation elicited by hypercapnia (arterial CO2 partial pressure = 62.7 +/- 3; n = 5) was preserved. In the rest of the brain the vasodilation elicited from FN was largely unaffected. We conclude that the vasodilation evoked from FN in cerebral cortex depends on the integrity of a restricted population of local neurons that interact with the local microvasculature.

A hierarchical organization of blood pressure during natural behaviour in rat and the effects of central catecholamine neurons thereon.

1. Changes in arterial pressure associated with a repertoire of natural behaviour patterns in the rat were examined. 2. A hierarchy of such changes was found. Eating and drinking were associated with higher pressures than grooming and exploration, which in turn were associated with higher pressures than resting. 3. Desynchronized sleep was associated with higher pressures than slow-wave sleep. 4. Lesions of the catecholamine neurons of the A2 region of the medulla did not disrupt the normal hierarchy but resulted in an exaggerated pressor response.