Cellular substrate of the histochemically defined striosome/matrix system of the caudate nucleus: a combined Golgi and immunocytochemical study in cat and ferret

In order to learn what morphological substrate might underly the histochemical compartments of the neostriatum, sections of the caudate nucleus and the putamen of cats and ferrets were stained immunocytochemically with antisera directed against several neuropeptides and transmitter-related enzymes and were then Golgi-impregnated. Adjacent sections were stained to reveal acetylcholinesterase activity to identify the acetylcholinesterase-poor striosomes. The immunostaining produced by several of the antibody preparations was in register with the acetylcholinesterase-poor striosomes but the most prominent staining of these zones occurred with the antibodies directed against substance P. The striosomes were delineated by intense substance P-immunostaining of neuronal perikarya and dendrites, and in the rostral and dorsal caudate nucleus the boundary between substance P-immunostained and extrastriosomal matrix was abrupt. For these reasons we analysed Golgi-impregnated neurons in sections immunostained for substance P in order to assess the influence of the chemically defined striosomal architecture on the position and dendritic arborization of neurons located both within the striosomes and within the extrastriosomal matrix. The most commonly impregnated neurons were of the medium-size densely spiny class. Those that were present within the striosomes and lay within one dendritic radius of the boundary were divided into two types: (1) neurons whose dendritic arborization was apparently not influenced by the boundary and (2) neurons whose dendritic arborization was markedly influenced by the boundary. For neurons of the latter type, dendrites either emerged from the parts of the perikaryon away from the boundary, so avoiding crossing it, or they exhibited abrupt changes in their course, apparently to avoid crossing the boundary. Spiny neurons located in the extrastriosomal matrix but close to the striosomal boundary had dendrites that were either influenced by, or not influenced by the compartmental boundary. We conclude that there is a specific cytoarchitecture underlying the histochemical compartments of the neostriatum and that different sub-populations of medium-size spiny neurons underly (1) the segregation of information flow in striosomes and the extrastriosomal matrix and (2) communication between striosomes and the extrastriosomal matrix.     

Glutamate-like immunoreactivity in neurons of the laterodorsal tegmental and pedunculopontine nuclei in the rat

Studies of the pedunculopontine (PPT) and laterodorsal tegmental (LDT) nuclei in the mesopontine tegmentum have emphasized the organization and projections of the cholinergic neurons. We report here that exhibiting glutamate immunoreactivity are present in both the LDT and PPT. These glutamatergic neurons are interspersed among the cholinergic neurons within both nuclei with no apparent segregation. These data raise the possibility that excitatory amino acids contribute to the functions of the LDT and PPT.     

Physicochemical, mechanical, and biological properties of commercial membranes for GTR.

Barrier membranes for guided tissue regeneration (GTR) to treat bone defects have to satisfy criteria of biocompatibility, cell-occlusiveness, spacemaking, tissue integration, and clinical manageability. In this study, the morphological and mechanical properties of two commercial biodegradable membranes (Resolut LT and Biofix) as a function of the incubation time have been compared. Moreover, their permeability to both fluids and epithelial cells as well as the bacteria adhesion have been evaluated. The membranes are asymmetric and composed of a dense polymeric layer coupled with nonwoven (Resolut LT) or woven (Biofix) fibers. Both of the membranes, when incubated in complete culture medium, completely lose the structural and mechanical properties within 30 days. Moreover the results of solute permeability show that Resolut LT and Biofix membranes cannot be considered selective membranes to the solute crossing. On the contrary, they act as a barrier to the passage of the gingivial cells and to S. mutans bacteria.     

Spatial segregation within the sacral parasympathetic nucleus of neurons innervating the bladder or the penis of the rat as revealed by three-dimensional reconstruction

The purpose of the present investigations was (1) to examine the spatial organization of preganglionic neurons of the sacral parasympathetic nucleus in the lumbosacral spinal cord of male adult rats and (2) to search, in this nucleus, for a possible segregation of sub-populations of neurons innervating the penis or the bladder, respectively. To estimate their spatial organization, neurons of the sacral parasympathetic nucleus were retrogradely labeled by wheat germ agglutinin coupled to horseradish peroxidase applied to the central end of the sectioned pelvic nerve. The sub-populations of lumbosacral neurons innervating the corpus cavernosum of the penis or the dome of the bladder were identified using transsynaptic retrograde labeling by pseudorabies virus injected into these organs in different rats. In both wheat germ agglutinin-labeled and pseudorabies virus-labeled rats, serial coronal sections were cut through the spinal L5-S1 segments. Labeled neurons were revealed by histochemistry (peroxidase experiments) or immunohistochemistry (pseudorabies virus experiments). By means of a three-dimensional reconstruction software developed in our laboratory, three-dimensional models were calculated from each spinal section image series. They revealed the spatial organization of (i) preganglionic neurons and (ii) neurons innervating the bladder or the penis. The different three-dimensional models were subsequently merged into a single one which revealed the segregation, within the sacral parasympathetic nucleus, of the sub-populations of neurons. Neurons labeled by virus injected into the penis extended predominantly from the rostral part of the L6 segment to the rostral part of the S1 segment while those labeled by bladder injections were distributed predominantly from the caudal part of the L6 segment to the caudal part of the S1 segment.These results support the hypothesis of a viscerotopic organization of sacral neurons providing the spinal control of pelvic organs.     

Encoding of electrical, thermal, and mechanical noxious stimuli by subnucleus reticularis dorsalis neurons in the rat medulla

1. In anesthetized rats, recordings were made within the medullary subnucleus reticularis dorsalis (SRD) from neurons that exhibited convergence of nociceptive inputs from the entire body. Neurons with total nociceptive convergence (TNC) responded to suprathreshold percutaneous electrical stimuli (2-ms duration) with an early and a late peak due to activation of A delta- and C-fibers, respectively, no matter which part of the body was stimulated. Neurons with partial nociceptive convergence (PNC) responded to the same stimuli with an A delta-peak regardless of which part of the body was stimulated and with a C-peak of activation from some, mainly contralateral, parts of the body. The characteristics of the responses of these neurons to the application of graded intensities of electrical, thermal, and mechanical stimuli were analyzed. 2. All TNC neurons and 85% of PNC neurons responded to A delta- and C-fiber activation following percutaneous electrical stimulation of the contralateral hindpaw. With regard to A delta-fiber-evoked responses, a linear relationship between the logarithm of the applied current and the magnitude of the responses was found within the 0.25- to 6.0-mA and 0.5- to 24-mA ranges for TNC and PNC neurons, respectively; however, these curves were essentially similar. With regard to C-fiber-evoked responses, such a linear relationship was found within the 1.5- to 6.0-mA range for both types of SRD neurons, although the TNC neurons presented larger C-fiber-evoked responses than did the PNC neurons. 3. TNC and PNC neurons linearly increased their discharges during the application of noxious thermal stimuli to the contralateral hindpaw within the range 44-52 degrees C; the mean responses evoked by noxious heat from TNC neurons were of higher magnitude than those from PNC neurons. The majority of SRD neurons presented long-lasting afterdischarges, especially with the highest temperature employed (52 degrees C). 4. TNC neurons monotonically increased their discharges during graded mechanical or thermal stimulation of the tail. When mechanical stimuli were applied, a linear relationship was found between the logarithm of the strength of the mechanical stimulus and the neuronal discharges, in the 5.3- to 7.4-N/cm2 range. With thermal stimulation, TNC neurons linearly increased their discharges in the 44-52 degrees C range. When increasing amounts of the tail were immersed in a 50 degrees C waterbath, TNC neurons increased their discharges within a restricted range of tail surface areas (0.9-5.7 cm2); further increases in the stimulated surface size were not followed by increases in firing rate.(ABSTRACT TRUNCATED AT 400 WORDS)     

Concept and computational design for a bioartificial nephron-on-a-chip

A MEMS-based, (Micro Electro Mechanical System) bioartificial device is proposed for replicating the function of a single nephron. Consistent with the anatomy and physiology of humans, our device has 3 distinct sections, replicating the function of the glomerulus, the proximal tubule, and the loop of Henle. Construction of a bioartificial loop of Henle in particular requires control of diffusion-scale features. The proposed device can be built using existing microfabrication technologies and populated with various renal cell types. A computational model is also developed to analyze the coupled, multiphase mass transport in this system. Using the model, a design is generated with flow and solute transport properties matching those of the human nephron.     

Neurons containing retrogradely transported Fluoro-Gold exhibit a variety of lysosomal profiles: A combined brightfield, fluorescence, and electron microscopic study

Summary The advantages of axonally transported Fluoro-Gold as a retrograde fluorescent marker are numerous. The objective of the present study was to determine whether transported Fluoro-Gold is visible in either semi-thin sections for light microscopy or thin sections for electron microscopy. Rats received injections of Fluoro-Gold into either the striatum or thoracic spinal cord. After appropriate survival times, labelled neurons were observed with the fluorescence microscope in brain regions that are known to project to the injected areas.Sections that contained labelled cells were embedded in plastic and examined with a fluorescence microscope. Semi-thin sections of unosmicated tissue displayed high-resolution fluorescent labelling of somata and dendrites. In contrast, osmicated tissue did not fluoresce, but numerous dark granules were observed in the dendritic and perikaryal cytoplasm of labelled neurons in toluidine blue stained sections that were examined with brightfield optics. The unosmicated tissue did not display these granules, and this finding suggested that the granules are composed of membranes. Neurons in other brain regions that are known not to project to the injection sites did not contain these dark granules.Adjacent thin sections examined with the electron microscope displayed numerous electron-dense, lysosome-like organelles in the cytoplasm of labelled neurons. The electron density of these organelles was greater than that of lysosomes in unlabelled neurons. Three types of distinctive organelles were observed in these preparations: (1) relatively dense concentric lamellar bodies of various sizes; (2) heterogeneous or lipofuscin-like lysosomes; and (3) coarse grained lysosomes. Control sections and unlabelled neurons did not display these organelles. Therefore, these organelles appear to correlate with Fluoro-Gold localized within the somata and dendrites of retrogradely labelled neurons. It is not known if they are the Fluoro-Gold itself, or represent a physiological effect on membranes. The results of this study indicate that Fluoro-Gold may be useful for tract tracing at the electron microscopic level.     

The glutamate decarboxylase-, leucine enkephalin-, methionine enkephalin- and substance P-immunoreactive neurons in the neostriatum of the rat and cat: evidence for partial population overlap

We have examined the populations of neurons in the neostriatum of both rat and cat that are immunoreactive for glutamate decarboxylase, [Leu]enkephalin, [Met]enkephalin and substance P. Neurons that were immunoreactive for glutamate decarboxylase made up 47% of the neurons in our samples from the rat and ranged from 39 to 49% of the neurons in the cat. Those immunoreactive for [Leu]enkephalin made up 44-49% of the neurons in rat neostriatum, and 38-47% in the cat, and those immunoreactive for [Met]enkephalin made up 36-41% of the neurons in rat and 43-49% of the neurons in the cat. Substance P-immunoreactive neurons made up 30-38% of neurons in rat and 32-39% in cat. Most substance P neurons (particularly the most darkly staining ones) were, however, clustered such that they were most numerous in the patch compartment of neostriatum; within the patches the substance P neurons comprised 59% of neurons in the rat and 55% in cat, but in the matrix substance P neurons comprised only 32% of neurons in the rat and 25% in the cat. Samples taken from sections processed for two-color double labeling immunocytochemistry revealed that 12% of neurons label for both glutamate decarboxylase and [Leu]enkephalin, 12% for both glutamate decarboxylase and [Met]enkephalin, 11-12% for both glutamate decarboxylase and substance P, and 17% for both [Met]enkephalin and substance P. These results provide evidence for chemical heterogeneity within the medium-sized neostriatal neurons, and provide the first evidence for coexistence of glutamate decarboxylase and substance P within a single neuron, and the first evidence for the coexistence for substance P and [Met]enkephalin within single neurons of the central nervous system.     

Neurotransmitter segregation: functional and plastic implications

Synaptic cotransmission is the ability of neurons to use more than one transmitter to convey synaptic signals. Cotransmission was originally described as the presence of a classic transmitter, which conveys main signal, along one or more cotransmitters that modulate transmission, later on, it was found cotransmission of classic transmitters. It has been generally accepted that neurons store and release the same set of transmitters in all their synaptic processes. However, some findings that show axon endings of individual neurons storing and releasing different sets of transmitters, are not in accordance with this assumption, and give support to the hypothesis that neurons can segregate transmitters to different synapses. Here, we review the studies showing segregation of transmitters in invertebrate and mammalian central nervous system neurons, and correlate them with our results obtained in sympathetic neurons. Our data show that these neurons segregate even classic transmitters to separated axons. Based on our data we suggest that segregation is a plastic phenomenon and responds to functional synaptic requirements, and to 'environmental' cues such as neurotrophins. We propose that neurons have the machinery to guide the different molecules required in synaptic transmission through axons and sort them to different axon endings. We believe that transmitter segregation improves neuron interactions during cotransmission and gives them selective and better control of synaptic plasticity.     

Transient co-localization of calretinin, parvalbumin, and calbindin-D28k in developing visual cortex of monkey

Summary This paper reports a double-labelling immunocytochemical study of the three calcium-binding proteins calretinin, parvalbumin, and calbindin-D28k in developing and adultMacaca primary visual cortex. In adult visual cortex, each protein marks a subset of GABAergic neurons with a characteristic laminar distribution and virtually no co-localization was found between these three proteins, suggesting that each calcium-binding protein may serve as a marker for one or more cortical subcircuits. The immature visual cortex, immunostained using identical techniques was then analysed to determine if each calcium-binding protein could serve as a developmental marker for these circuits. The Cajal-Retzius cells of layer 1 contained all three proteins during development. Calbindin-D28k and calretinin were co-localized starting at Fd (foetal day) 45 and after Fdl25, parvalbumin also was present in the same Cajal-Retzius cells. All three proteins continued to be expressed until the Cajal-Retzius disappeared postnatally. In layers 2–6 calbindin-D28k and calretinin were never co-localized. In contrast, parvalbumin and calretinin were found in neurons of deep layer 3 from Fd 155 to postnatal (P6) weeks with a few persisting even later. Before birth almost all PV⁺ neurons in layers 4–6 were CaB⁺, but by P3 weeks only a few PV⁺/CaB⁺ neurons remained in layer 4C and these completely disappeared by P6 weeks. Co-localization in layer 4 neurons overlaps the period of ocular dominance segregation, suggesting that the onset of cortical maturity coincides with segregation of calcium-binding proteins within the GABA interneurons.     

The raccoon lateral cervical nucleus: mediolateral organization of GABA-positive and GABA-negative neurons and fibers

In the lateral cervical nucleus (LCN) of the cat, GABA-immunoreactive neurons and substance P-immunoreactive fibers are concentrated in the medial part of the nucleus, whereas in the monkey LCN no preferential locations have been identified. In raccoons, substance P-immunoreactive fibers display a distribution pattern similar to that in cats. However, the presence and distribution of GABA-immunoreactive neurons in the raccoon LCN has not been examined, and it is therefore not known whether raccoons are similar to cats or primates in this respect. Thus, in the present study, the raccoon LCN was examined for the presence and distribution of GABA-immunoreactive cells with respect to their numbers, locations, and sizes. The distribution of GABA-positive fibers and varicosities within the LCN was also investigated. The results of measurements of cross-sectional areas of LCN neurons indicate a trend toward decreasing cell size along the dorsolateral to medial axis of the raccoon LCN. Compared to neurons of the centrally located ventromedial division, neurons are statistically significantly larger in the dorsolateral division and smaller in the medial division of the nucleus. Cell counts in post-embedding-stained semithin sections through the nucleus revealed an average of 8,700 neurons per LCN. Approximately 4% of LCN neurons are GABA-immunoreactive. These neurons are small and most (80%) of them are located in the medial third of the LCN. In contrast, GABA-immunoreactive fibers and varicosities are present in about equal density throughout the raccoon LCN. Thus, the distributions of GABA-immunoreactive neurons and neuron sizes in the raccoon LCN conform closely to those in cats. Together with previous observations in cats and raccoons, the present findings support the notion that these small GABA-immuno-reactive neurons may be local circuit inhibitory neurons and indicate the presence of a mediolateral segregation that may be of fundamental importance for the functional organization of the carnivore LCN.     

The rat perirhinal cortex: A review of anatomy, physiology, plasticity, and function

The perirhinal cortex is located in a pivotal position to influence the flow of information into and out of the hippocampal formation. In this review, we examine the anatomical, physiological and functional properties of the rat perirhinal cortex. Firstly, we review the properties of the perirhinal cortex itself, we describe how it can be separated into two distinct subregions and consider how it differs from other neighbouring regions in terms of cell type, cellular organisation and its afferent and efferent projections. We review the forms of neurotransmission present in the perirhinal cortex and the morphological, electrophysiological and plastic properties of its neurons. Secondly, we review the perirhinal cortex in the context of its connections with other brain areas; focussing on the projections to cortical, subcortical and hippocampal/parahippocampal regions. Particular attention is paid the anatomical and electrophysiological properties of these projections. Thirdly, we review the main functions of the perirhinal cortex; its roles in perception, recognition memory, spatial and contextual memory and fear conditioning are explored. Finally, we discuss the idea of anatomical, electrophysiological and functional segregation within the perirhinal cortex itself and as part of a hippocampal-parahippocampal network and suggest that understanding this segregation is of critical importance in understanding the role and contributions made by the perirhinal cortex in general.     

Somatostatin and vasoactive intestinal polypeptide-like immunoreactive cells and terminals in the retrohippocampal region of the rat brain

The retrohippocampal region of the rat brain was analyzed by using immunohistochemistry with specific antibodies against somatostatin (SOM) and vasoactive intestinal polypeptide (VIP). Specifically immunoreactive neurons and terminal processes were labeled with either the anti-SOM or anti-VIP antiserum and they were referred to as SOM-like immunoreactive (SOM-LI) or VIP-like immunoreactive (VIP-LI) neurons and processes, respectively. The retrohippocampal region was rich in neuronal cell bodies and terminal processes showing immunoreactivity for SOM and VIP. In the entorhinal area SOM-LI neurons were located mainly in layers IV through VI and the VIP-LI neurons were found mainly in layers I through III. Thick (70-120 microns) sections treated with the immunoperoxidase method to achieve a Golgi-like staining pattern showed that cytological differences existed between SOM- and VIP-positive neurons. SOM-LI neurons were usually multipolar, fusiform, or occasionally pyramidal while VIP-LI neurons were usually bipolar, stellate, or fusiform. SOM-LI and VIP-LI axons and preterminal processes were differentially distributed within the laminae of the retrohippocampal region. VIP-LI terminals were found throughout all layers except layer I. SOM-LI terminals were found primarily in the molecular layers of all areas, layer IV of the medical and lateral entorhinal areas, and in the angular bundle. Thus, SOM-LI and VIP-LI neurons are distinguished by their morphology and their different distribution within the cortical layers and areas of the retrohippocampal region.     

Dissociation of spatial-, object-, and sound-coding neurons in the mediodorsal nucleus of the primate thalamus

The mediodorsal nucleus (MD) is the thalamic gateway to the prefrontal cortex, an area of the brain associated with spatial and object working memory functions. We have recorded single-neuron activities from the MD nucleus in monkeys trained to perform spatial tasks with peripheral visual stimuli and a nonspatial task with foveally presented pictures of objects and faces-tasks identical to those we have previously used to map regional specializations in the dorso- and ventro-lateral prefrontal cortex, respectively. We found that MD neurons exhibited categorical specificity-either responding selectively to locations in the spatial tasks or preferentially to specific representations of faces and objects in the nonspatial task. Spatially tuned neurons were located in parts of the MD connected with the dorsolateral prefrontal cortex while neurons responding to the identity of stimuli mainly occupied more ventral positions in the nucleus that has its connections with the inferior prefrontal convexity. Neuronal responses to auditory stimuli were also examined, and vocalization sensitive neurons were found in more posterior portions of the MD. We conclude that MD neurons are dissociable by their spatial and nonspatial coding properties in line with their cortical connections and that the principle of information segregation in cortico-cortical pathways extends to the "association" nuclei of the thalamus.     

Cyclic Deformation-Induced Solute Transport in Tissue Scaffolds with Computer Designed,Interconnected, Pore Networks: Experiments and Simulations

Nutrient supply and waste removal in porous tissue engineering scaffolds decrease from the periphery to the center, leading to limited depth of ingrowth of new tissue into the scaffold. However, as many tissues experience cyclic physiological strains, this may provide a mechanism to enhance solute transport in vivo before vascularization of the scaffold. The hypothesis of this study was that pore cross-sectional geometry and interconnectivity are of major importance for the effectiveness of cyclic deformation-induced solute transport. Transparent elastic polyurethane scaffolds, with computer-programmed design of pore networks in the form of interconnected channels, were fabricated using a 3D printing and injection molding technique. The scaffold pores were loaded with a colored tracer for optical contrast, cyclically compressed with deformations of 10 and 15% of the original undeformed height at 1.0 Hz. Digital imaging was used to quantify the spatial distribution of the tracer concentration within the pores. Numerical simulations of a fluid–structure interaction model of deformation-induced solute transport were compared to the experimental data. The results of experiments and modeling agreed well and showed that pore interconnectivity heavily influences deformation-induced solute transport. Pore cross-sectional geometry appears to be of less relative importance in interconnected pore networks. Validated computer models of solute transport can be used to design optimal scaffold pore geometries that will enhance the convective transport of nutrients inside the scaffold and the removal of waste, thus improving the cell survivability deep inside the scaffold.     

Subcellular organization of GABAergic synapses: role of ankyrins and L1 cell adhesion molecules

In vertebrate nervous systems, different classes of synaptic inputs are often segregated into restricted compartments of target neurons. For example, distinct types of GABAergic interneurons preferentially innervate subcellular domains and have been implicated in the precise temporal regulation of integration within neurons and activity within networks. Recent studies suggest that the subcellular segregation of different classes of GABAergic synapses is largely genetically determined. The localization and signaling of L1 family immunoglobulin proteins recruited by ankyrin-based membrane adaptors might serve as compartmental labels, which contribute to subcellular synapse organization in cerebellar Purkinje neurons.     

Further observations on the relationship between adenosine deaminase-containing axons and trigeminal mesencephalic neurons: an electron microscopic, immunohistochemical and anterograde tracing study

The somas of primary afferent neurons in the mesencephalic nucleus of the trigeminal nerve in rat have a dense investment of axons immunoreactive for the enzyme adenosine deaminase. We previously suggested that these axons may originate from adenosine deaminase-immunoreactive neurons located in the tuberomammillary nucleus of the hypothalamus [Nagy et al. (1986) Neuroscience 17, 141-156]. Anterograde tracing and immunohistochemical techniques were used to investigate this possibility further. In addition, the appearance of adenosine-immunoreactive axons and the nature of their interactions with mesencephalic neurons was examined ultrastructurally. After injections of either Phaseolus vulgaris-leucoagglutinin or wheat germ agglutinin-horseradish peroxidase into the region of the tuberomammillary nucleus, punctate deposits of anterogradely transported tracer, detected by immunoperoxidase methods, were seen surrounding mesencephalic neurons. In sections immunostained for tracer and adenosine deaminase by double immunofluorescence, some fibres in the periaqueductal gray matter and around Mes V somas were found to be labelled for both the lectin and the enzyme. Ultrastructurally, only a single morphological class of adenosine deaminase-immunoreactive axons adjacent to, or indenting the cytoplasmic membranes of, large somas in the mesencephalic nucleus could be recognized; they were varicose and contained relatively large immunoreactive vesicles ranging in diameter from 45 to 70 nm. Occasionally, thin processes of these axons could be traced back to small adenosine deaminase-positive neuronal cell bodies located not within the tuberomammillary nucleus, but rather, within the periaqueductal gray matter. In serial ultrathin sections, membrane specializations resembling synaptic junctions were sometimes seen at points where mesencephalic somas were in contact with adenosine deaminase-immunoreactive terminals. Somas within the mesencephalic nucleus also formed such junctions with non-immunoreactive boutons which were morphologically different from, and often seen in close proximity to, those containing adenosine deaminase. These results indicate that in addition to possible afferents from the tuberomammillary nucleus, primary sensory somas within the mesencephalic nucleus are also associated with axonal processes originating from adenosine deaminase-positive neurons located within the periaqueductal gray matter. The infrequent synaptic contacts between these somas and adenosine deaminase-positive axons, despite their close anatomical arrangement, is suggestive of a diffuse endocrine or neurocrine type of axonal relationship with mesencephalic somas or with the n     

Separate populations of neurons within the paraventricular hypothalamic nucleus of the rat project to vagal and thoracic autonomic preganglionic levels and express c-Fos protein induced by lithium chloride

The role of different hypothalamic nuclei, particularly the paraventricular nucleus (PVN), in the control of food intake and feeding behaviour is well known. It is also well established that lithium chloride (LiCl) causes various disorders in feeding behaviour. In this study, we analyzed the precise distribution of hypothalamic neurons activated by i.p. LiCl administration (LCA neurons) and compared it to that of hypothalamic neurons which project to autonomic preganglionic levels (HAP neurons). We also analysed the possibility that some neurons belong to both populations of nerve cells. To this end, a multiple-labelling technique, using two retrograde fluorescent tracers together with c-Fos-like immunohistochemistry, was performed. Fast Blue was injected in the dorsal motor nucleus of the vagus and Fluorogold (FG) in the thoracic intermedial-lateral cell column, to trace parasympathetic and sympathetic pathways, respectively. LiCl was used as stimulus for c-Fos-like immunohistochemistry. HAP neurons were located mainly in the dorsal, ventral and lateral regions of the parvocellular PVN, while LCA neurons were observed predominantly in the magnocellular region of the PVN rostrally to HAP neurons. A significant number of FG/Fos double-labelled neurons were located in the dorsal parvocellular subnucleus of the PVN (dp) in the LiCl-stimulated rats. We concluded that there is a clear segregation of LCA neurons from HAP neurons within the PVN. The presence of FG/Fos double-labelled neurons in the dp suggests that this nucleus could mediate a sympathetic response after LiCl administration.     

Cannabinoid agonist, CP 55,940, prevents capsaicin-induced sensitization of spinal cord dorsal horn neurons

Low doses of cannabinoids applied intrathecally attenuate capsaicin-evoked heat and mechanical hyperalgesia via CB1 receptors. Although cannabinoids produce antinociception, in part, by attenuating responses of nociceptive neurons in the spinal cord, few studies have examined the effect of cannabinoids on sensitization of spinal neurons. We therefore investigated whether a cannabinoid receptor agonist, CP 55,940, attenuated excitation and sensitization of spinal nociceptive neurons produced by intraplantar injection of 0.1% capsaicin (10 microl). In rats, wide-dynamic-range (WDR) and high-threshold (HT) neurons were classified according to responses evoked by mechanical stimuli of varying intensity. CP 55,940 (10 microg in 50 microl) or vehicle was applied directly to the spinal cord and responses to mechanical (von Frey monofilament) and heat stimuli were recorded 10 min after drug treatment. CP 55,940 alone did not alter responses to mechanical stimuli; however the enhanced responses to mechanical stimuli after injection of capsaicin into the receptive field were dose dependently attenuated in both HT and WDR neurons. Vehicle-treated neurons increased their response to 300.6 +/- 52.1% of baseline after capsaicin, whereas CP 55,940-treated neurons responded at 153.0 +/- 27.1% of baseline. The effects of CP 55,940 on sensitization to heat were less pronounced; however, CP 55,940 attenuated the capsaicin-evoked decrease in heat threshold in HT neurons. The attenuation by CP 55,940 of sensitization to mechanical stimuli was blocked by pretreatment of the spinal cord with the CB1 receptor antagonist, SR141716A. These studies demonstrate that cannabinoid application to the spinal cord prevents central sensitization.     

Characterization of substance P- and [Met]enkephalin-immunoreactive neurons in the caudate nucleus of cat and ferret by a single section Golgi procedure

Modifications of the single-section Golgi-impregnation procedure of Gabbott and Somogyi are described. The modifications allow easier and more rapid preparation of the sections for Golgi-impregnation and easier handling of large numbers of serial sections. The technique consists of placing a section that has been treated with osmium tetroxide and potassium dichromate on a microscope slide and "sandwiching" it with a second microscope slide. The two slides are held together at one end by tape and the assembly is dipped into a solution of silver nitrate. Golgi-impregnation of neurons occurs within a few hours and is generally complete within 12 h. The technique has been applied to sections through the caudate nucleus of the cat and ferret in order to define the morphological characteristics of striatal substance P- and methionine enkephalin-immunoreactive neurons. Sections were first incubated to reveal the immunoreactive structures and then subjected to the Golgi method. Golgi-impregnated neurons that were immunoreactive for either substance P or methionine enkephalin had medium-size perikarya from which several dendrites emerged. The dendrites branched close to the perikaryon; secondary and higher order dendrites were densely laden with spines, as many as 15 spines per 10 microns of dendrite. It is concluded that both striatal substance P-containing and methionine enkephalin-containing neurons are of the medium-size densely spiny type. Medium-size densely spiny neurons may be homogeneous with respect to their somatodendritic morphology but heterogeneous with respect to their chemical characteristics and axonal morphology.