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V M Pickel

Publications and source records attributed to V M Pickel.

At least 91 records · Page 5Linked to original sources
PubMed1996

Ultrastructural localization of mu-opioid receptors in the superficial layers of the rat cervical spinal cord: extrasynaptic localization and proximity to Leu5-enkephalin.

Many of the analgesic effects of opiate drugs and of endogenous opioid ligands, such as Leu5-enkephalin (LE) are thought to be mediated in part by mu-opioid receptors (MOR) in the dorsal horn of the spinal cord. To establish the cellular sites for the spinally mediated analgesic effects of MOR activation and the potential anatomical substrates for interactions with LE, we examined the ultrastructural localization of MOR and LE immunoreactivities in the adult rat cervical spinal cord (C3-C5). Anti-MOR sera recognizing the carboxyl terminal domain of MOR was localized using immunoperoxidase and immunogold-silver methods. mu-opioid receptor-like immunoreactivity (MOR-LI) was observed mainly in the superficial layers of the dorsal horn. Electron microscopy of this region revealed that small unmyelinated axons and axon terminals constituted 48% (91/189) and 15% (28/189), respectively, while dendrites comprised 36% (68/189) of the total population of neuronal profiles containing the MOR. MOR-LI was localized mainly along extrasynaptic portions of the plasma membrane in both axons and dendrites. In sections dually labeled for MOR and LE, 21% (14/68) of the dendrites containing MOR-LI closely apposed or received synaptic contact from axon terminals exhibiting LE reaction product. The results provide the first ultrastructural evidence that within the dorsal horn of the spinal cord, LE, as well as exogenous opiates may alter both axonal release of neurotransmitters and postsynaptic responsiveness of target neurons to afferent input through activation of extrasynaptic MOR.

Amino Acid Sequence↗
PubMed1996

Ultrastructural evidence for prominent distribution of the mu-opioid receptor at extrasynaptic sites on noradrenergic dendrites in the rat nucleus locus coeruleus.

Physiological studies have indicated that agonists at the mu-opioid receptor (mu OR), such as morphine or the endogenous peptide methionine5-enkephalin, can markedly decrease the spontaneous activity of noradrenergic neurons in the locus coeruleus (LC). Messenger RNA and protein for mu OR are also densely expressed by LC neurons. During opiate withdrawal, increased discharge rates of LC neurons coincide with the expression of behavioral features associated with the opiate withdrawal syndrome. To better define the cellular sites for the physiological activation of mu OR in the LC and its relation to afferent terminals, we examined the ultrastructural localization of mu OR immunoreactivity in sections dually labeled for the catecholamine-synthesizing enzyme tyrosine hydroxylase (TH). Immunogold-silver labeling for mu OR (i-mu OR) was localized to parasynaptic and extrasynaptic portions of the plasma membranes of perikarya and dendrites, many of which also contained immunolabeling for TH. The dendrites containing exclusively i-mu OR were more numerous in the rostral pole of the LC. The i-mu OR in dendrites with and without detectable TH immunoreactivity were usually postsynaptic to unlabeled axon terminals containing heterogeneous types of synaptic vesicles and forming asymmetric synaptic specializations characteristic of excitatory-type synapses. These results provide the first direct ultrastructural evidence that mu OR is strategically localized to modulate the postsynaptic excitatory responses of catecholamine-containing neurons in the LC.

Animals↗
PubMed1996

Immunolabeling of Mu opioid receptors in the rat nucleus of the solitary tract: extrasynaptic plasmalemmal localization and association with Leu5-enkephalin.

Activation of the mu opioid receptor (MOR) by morphine within the caudal nucleus of the solitary tract (NTS) is known to mediate both cardiorespiratory and gastrointestinal responses. Leu5-enkephalin (LE), a potential endogenous ligand for MOR, is also present within neurons in this region. To determine the cellular sites for the visceral effects of MOR ligands, including LE, we used immunogold-silver and immunoperoxidase methods for light and electron microscopic localization of antisera against MOR (carboxyl terminal domain) and LE in the caudal NTS of rat brain. Light microscopy of coronal sections through the NTS at the level of the area postrema showed MOR-like immunoreactivity (MOR-LI) and LE labeling in punctate processes located within the subpostremal, dorsomedial and medial subnuclei. Electron microscopy of sections through the medial NTS at this level showed gold-silver particles identifying MOR-LI prominently distributed to the cytoplasmic side of the plasma membranes of axons and terminals. MOR labeled terminals formed mostly symmetric (inhibitory-type) synapses but sometimes showed multiple asymmetric junctions, characteristic of excitatory visceral afferents. MOR-LI was also present along extrasynaptic plasma membranes of dendrites receiving afferent input from unlabeled and LE-labeled terminals. We conclude that MOR ligands, possibly including LE, can act at extrasynaptic MORs on the plasma membranes of axons and dendrites in the caudal NTS to modulate the presynaptic release and postsynaptic responses of neurons. These are likely to include local inhibitory neurons and both gastric and cardiorespiratory afferents known to terminate in the subnuclei with the most intense MOR-LI.

Amino Acid Sequence↗
PubMed1996

Ultrastructural immunocytochemical localization of neurotensin and the dopamine D2 receptor in the rat nucleus accumbens.

The neuroleptic-like effects of neurotensin (NT) are thought to be due to interactions with dopamine (DA) acting primarily at D2 receptors within the nucleus accumbens septi (Acb). Using electron microscopic dual labeling immunocytochemistry, we sought to demonstrate cellular substrates for functional interactions involving NT and DA D2 receptors in the adult rat Acb. Peroxidase reaction product representing D2 receptor-like immunoreactivity (D2-LI) was seen along membranes of Golgi lamellae and multivesicular bodies of perikarya containing immunogold labeling representing NT-LI. Dually labeled somata usually contained highly indented nuclei, a characteristic of aspiny neurons. Dendrites also occasionally colocalized the two immunomarkers. Other somata, dendrites, and all axon terminals were singly labeled with either NT-LI or D2-LI. In distinct sets of terminals, NT-LI was commonly associated with large, dense-cored vesicles, whereas D2-LI was found along the plasmalemma and over nearby small clear vesicles. Each type of terminal comprised approximately 20% of synaptic input to NT-immunoreactive dendrites. Similar proportions of terminals containing NT-LI or D2-LI contacted unlabeled (approximately 55%) or NT-labeled (approximately 35%) dendrites and, occasionally, were observed converging onto common dendrites. Terminals containing NT-LI or D2-LI also were often closely apposed. These findings provide the first ultrastructural evidence that: (1) NT and D2 receptors are colocalized in aspiny neurons and dendrites, (2) NT may produce a direct postsynaptic effect on neurons receiving input from terminals which are presynaptically modulated by DA via D2 receptors, and (3) NT and DA acting at D2 receptors may interact through presynaptic modulation of common axon terminals.

Animals↗
PubMed1996

Input from central nucleus of the amygdala efferents to pericoerulear dendrites, some of which contain tyrosine hydroxylase immunoreactivity.

Light microscopic anterograde tracing studies indicate that neurons in the central nucleus of the amygdala (CNA) project to a region of the dorsal pontine tegmentum ventral to the superior cerebellar peduncle which contains noradrenergic dendrites of the nucleus locus coeruleus (LC). However, it has not been established whether the efferent terminals from the CNA target catecholamine-containing dendrites of the LC or dendrites of neurons from neighboring nuclei which may extend into this region. To examine this question, we combined immunoperoxidase labeling of the anterograde tracer biotinylated dextran amine (BDA) from the CNA with immunogold-silver labeling of the catecholamine-synthesizing enzyme tryrosine hydroxylase (TH) in the rostrolateral LC region of adult rats. By light microscopy, BDA-labeled processes were dense in the dorsal pons within the parabrachial nuclei as well as in the pericoerulear region immediately ventral to the superior cerebellar peduncle. Higher magnification revealed that BDA-labeled varicose fibers overlapped TH-labeled processes in this pericoerulear region. By electron microscopy, anterogradely labeled axon terminals contained small, clear as well as some large dense core vesicles and were commonly apposed to astrocytic processes along some portion of their plasmalemma. BDA-labeled terminals mainly formed symmetric type synaptic contacts characteristic of inhibitory transmitters. Of 250 BDA-labeled axon terminals examined where TH immunoreactivity was present in the neuropil, 81% contacted unlabeled and 19% contacted TH-labeled dendrites. Additionally, amygdala efferents were often apposed to unlabeled axon terminals forming asymmetric (excitatory type) synapses. These results demonstrate that amygdaloid efferents may directly alter the activity of catecholaminergic and non-catecholaminergic neurons in this pericoerulear region of the rat brain. Furthermore, our study suggests that CNA efferents may indirectly affect the activity of pericoerulear neurons through modulation of excitatory afferents. Amygdaloid projections to noradrenergic neurons may help integrate behavioral and visceral responses to threatening stimuli by influencing the widespread noradrenergic projections from the LC.

Amygdala↗
PubMed1996

Pre- and postsynaptic sites for serotonin modulation of GABA-containing neurons in the shell region of the rat nucleus accumbens.

The shell of the nucleus accumbens received a dense serotonergic innervation and contains abundant gamma-aminobutyric acid (GABA)-immunoreactive neurons. Moreover, serotonin (5-hydroxytryptamine: 5-HT) and GABA have been implicated in a variety of common motivational and motor-related functions partially ascribed in this brain area. We used immunoelectron microscopy of antisera directed against 5-HT and GABA in the same section of tissue to examine whether there were cellular substrates that might indicate more specific sites for functional interactions involving these transmitters in the shell region of the rat nucleus accumbens. Immunogold-silver labeling for GABA was localized to perikarya, dendrites, axons and axon terminals, whereas immunoperoxidase labeling for 5-HT was restricted to axons and axon terminals. Approximately half (187/366) of the 5-HT-immunoreactive axon terminals apposed or formed synaptic junctions with postsynaptic neurons. These junctions were mainly of the symmetric-type (83/187) characteristic of inhibitory transmitters, and were equally prevalent on dendrites with and without detectable gold-silver labeling for GABA. Of the 187 5-HT-labeled axon terminals with recognized synaptic contacts, 36% also showed convergence on a common dendrite with a GABA-labeled axon terminal. In addition, 5-HT- and GABA-immunoreactive axon terminals were commonly (83/366) identified in direct apposition to one another. Within a single plane of section, 41% of the apposed GABA-immunoreactive axon terminals formed symmetric-type junctions with dendrites or somata, whereas, the apposed 5-HT-labeled axon terminals rarely showed postsynaptic contacts. These results indicate that 5-HT-containing axon terminals may postsynaptically inhibit GABAergic neurons and their targets within the shell of the rat nucleus accumbens. Additionally, our results strongly suggest that, in this brain region, appositions between 5-HT and GABA axons and axon terminals may facilitate presynaptic interactions between these transmitter systems.

Animals↗
PubMed1996

Ultrastructural localization of the vesicular monoamine transporter-2 in midbrain dopaminergic neurons: potential sites for somatodendritic storage and release of dopamine.

Midbrain dopaminergic neurons are known to release dopamine from somata and/or dendrites located in the substantia nigra (SN) and the ventral tegmental area (VTA). There is considerable controversy, however, about the subcellular sites for somatodendritic dopamine storage in these regions. In the present study, we used dual-labeling electron microscopic immunocytochemistry to localize the vesicular monoamine transporter-2 (VMAT2), a novel marker for sites of intracellular monoamine storage, within identified dopaminergic (tyrosine hydroxylase-containing) neurons in the rat SN and VTA. In dopaminergic perikarya, immunogold labeling for VMAT2 was localized to the Golgi apparatus, tubulovesicles that resembled smooth endoplasmic reticulum (SER), and the limiting membranes of multivesicular bodies. In dopaminergic dendrites, VMAT2 was extensively localized to tubulovesicles that resembled saccules of SER, and less frequently localized to isolated small synaptic vesicles (SSVs) or large dense-core vesicles (DCVs). In rare cases, VMAT2-immunoreactive SSVs were clustered within the cytoplasm of an SN or a VTA dendrite. Dopaminergic dendrites in the VTA contained a significantly higher number of immunogold particles for VMAT2 per unit than those in the SN. Together, these observations support the proposal that dopamine is stored in and may be released from dendritic SSVs and DCVs, but suggest that the SER is the major site of dopamine storage within midbrain dopaminergic neurons. In addition, they provide new evidence that dopaminergic dendrites in the VTA may have greater potential for reserpine-sensitive storage and release of dopamine than those in the SN.

Animals↗
PubMed1996

Ultrastructural immunocytochemical localization of mu-opioid receptors in rat nucleus accumbens: extrasynaptic plasmalemmal distribution and association with Leu5-enkephalin.

mu-Opioid receptors and their endogenous ligands, including Leu5-enkephalin (LE), are distributed abundantly in the nucleus accumbens (NAC), a region implicated in mechanisms of opiate reinforcement. We used immunoperoxidase and/or immunogold-silver methods to define ultrastructural sites for functions ascribed to mu-opioid receptors and potential sites for activation by LE in the NAC. An antipeptide antibody raised against an 18 amino acid sequence of the cloned mu-opioid receptor (MOR) C terminus showed that MOR-like immunoreactivity (MOR-LI) was localized predominantly to extrasynaptic sites along neuronal plasma membranes. The majority of neuronal profiles containing MOR-LI were dendrites and dendritic spines. The dendritic plasma membranes immunolabeled for MOR were near sites of synaptic input from LE-labeled terminals and other unlabeled terminals forming either inhibitory or excitatory type synapses. Unmyelinated axons and axon terminals were also intensely but less frequently immunoreactive for MOR. Observed sites for potential axonal associations with LE included coexistence of MOR and LE within the same terminal, as well as close appositions between differentially labeled axons. Astrocytic processes rarely contained detectable MOR-LI, but also were sometimes observed in apposition to LE-labeled terminals. We conclude that in the rat NAC, MOR is localized prominently to extrasynaptic neuronal and more rarely to glial plasma membranes that are readily accessible to released LE and possibly other opioid peptides and opiate drugs. The close affiliation of MOR with spines receiving excitatory synapses and dendrites receiving inhibitory synapses provides the first direct morphological evidence that MOR selectively modulates postsynaptic responses to cortical and other afferents.

Animals↗
PubMed1996

GABAergic neurons in rat nuclei of solitary tracts receive inhibitory-type synapses from amygdaloid efferents lacking detectable GABA-immunoreactivity.

Gamma-aminobutyric acid (GABA) is a prominent inhibitory transmitter in both the central nucleus of the amygdala (Ce) and the medial nuclei of the solitary tracts (mNTS). These regions are reciprocally connected by anatomical pathways mediating the coordinated visceral responses to emotional stress. To further determine whether GABA is present in the amygdaloid efferents or their targets in the mNTS, we combined peroxidase labeling of Phaseolus vulgaris leucoagglutinin (PHA-L) or biotinylated dextran amine (BDA) anterogradely transported from the Ce with immunogold-silver detection of antibodies against GABA in the rat mNTS. By light microscopy, peroxidase labeling for either PHA-L or BDA was seen in varicose processes, whereas immunogold-silver labeling for GABA was detected in perikarya and processes throughout the rostrocaudal mNTS. The intermediate mNTS at the level of the area postrema, a region receiving mainly cardiorespiratory and gastric visceral afferents, were examined by electron microscopy. In this region, anterograde labeling was observed exclusively in unmyelinated axons and axon terminals. These terminals lacked detectable GABA-immunoreactivity, but formed symmetric synapses that are associated with inhibition. The targets of the anterogradely labeled terminals were medium-sized dendrites both with and without GABA-labeling. These dendrites often also received convergent input from terminals that were intensely GABA-immunoreactive. We conclude that visceral activation accompanying emotional response to stress is likely to involve inhibition of GABAergic neurons in the mNTS by non-GABA-containing amygdaloid efferents. Furthermore, our results indicate that the inhibition of these GABAergic neurons may be further augmented by release of GABA from other converging terminals in the mNTS.

Amygdala↗
PubMed1996

The dopamine transporter is localized to dendritic and axonal plasma membranes of nigrostriatal dopaminergic neurons.

Nigrostriatal dopaminergic neurons play an essential role in the central regulation of motor functions. These functions are initiated through the release of dopamine from axon terminals in the striatum or from dendrites in the substantia nigra (SN) and are terminated by the reuptake of dopamine by the sodium- and chloride-dependent dopamine transporter (DAT). DAT also can transport dopamine neurotoxins and has been implicated in the selective vulnerability of nigrostriatal dopaminergic neurons in major models of Parkinson's disease. We have used electron microscopic immunocytochemistry with an N-terminal domain anti-peptide antibody to examine the subcellular distribution of DAT in the rat SN and dorsolateral striatum. In the SN, immunogold labeling for DAT was localized to cytoplasmic surfaces of plasma membranes and smooth endoplasmic reticulum of dendrites and dendritic spines, few of which contained synaptic vesicles. Neuronal perikarya in the SN contained immunogold-labeled pleomorphic electron-lucent tubulovesicles but showed immunolabeling of plasma membranes only rarely. Axon terminals in the striatum contained extensive immunogold labeling of cytoplasmic surfaces of plasma membranes near aggregates of synaptic vesicles and less frequent labeling of intervaricose segments of plasma membrane or small electron-lucent vesicles. In sections dually labeled for DAT and the catecholamine-synthesizing enzyme tyrosine hydroxylase, both markers were colocalized in most profiles in the SN and striatum. These findings support the proposed topological model for DAT and suggest that this transporter is strategically located to facilitate uptake of dopamine and neurotoxins into distal dendritic and axonal processes of nigrostriatal dopaminergic neurons.

Amino Acid Sequence↗
PubMed1996

Pyramidal neurons in rat prefrontal cortex show a complex synaptic response to single electrical stimulation of the locus coeruleus region: evidence for antidromic activation and GABAergic inhibition using in vivo intracellular recording and electron microscopy.

Cognition and acquisition of novel motor skills and responses to emotional stimuli are thought to involve complex networking between pyramidal and local GABAergic neurons in the prefrontal cortex. There is increasing evidence for the involvement of cortical norepinephrine (NE) deriving from the nucleus locus coeruleus (LC) in these processes, with possible reciprocal influence via descending projections from the prefrontal cortex to the region of the LC. We used in vivo intracellular recording in rat prefrontal cortex to determine the synaptic responses of individual neurons to single electrical stimulation of the mesencephalic region including the nucleus LC. The most common response consisted of a late-IPSP alone or preceded by an EPSP. The presence of an early-IPSP following the EPSP was sometimes detected. Analysis of the voltage dependence revealed that the late-IPSP and early-IPSP were putative K(+)- and Cl- dependent, respectively. Synaptic events occurred following short delays and were inconsistent with the previously reported time for electrical activation of unmyelinated LC fibers. Moreover, systemic injection of the adrenergic antagonists propranolol (beta receptors), or prazosin (alpha 1 receptors), did not block synaptic responses to stimulation of the LC region. Finally, certain neurons were antidromically activated following electrical stimulation of this region of the dorsal pontine tegmentum. Taken together, these results suggest that the complex synaptic events in pyramidal neurons of the prefrontal cortex that are elicited by single electrical stimulation of the LC area are mainly due to antidromic activation of cortical efferents. Further insight into the chemical circuitry underlying these complex synaptic responses was provided by electron microscopic immunocytochemical analysis of the relations between the physiologically characterized neurons and either 1) GABA or 2) dopamine-beta-hydroxylase (DBH), a marker for noradrenergic terminals. GABA-immunoreactive terminals formed numerous direct symmetric synapses on somata and dendrites of pyramidal cells recorded and filled with lucifer yellow (LY). In contrast, in single sections, noradrenergic terminals immunoreactive for DBH rarely contacted LY-filled somata and dendrites. These results support the conclusion that IPSPs observed following single electrical stimulation of the LC region are mediated by GABA, with little involvement of NE. These IPSPs, arising from antidromic invasion of mPFC cells innervating the LC, may improve the signal-to-noise ratio and favor a better responsiveness of neighboring neurons to NE released in the mPFC.

Animals↗
PubMed1996

Enkephalin terminals form inhibitory-type synapses on neurons in the rat nucleus locus coeruleus that project to the medial prefrontal cortex.

Norepinephrine-containing fibres in the medial prefrontal cortex derive from the locus coeruleus, a brainstem nucleus which also receives a dense innervation of enkephalin-immunoreactive axon terminals. We combined immunogold-silver labelling of retrogradely transported FluoroGold from the medial prefrontal cortex with immunoperoxidase detection of leucine5-enkephalin in the same section of tissue through the locus coeruleus of adult rats. This dual-labelling experiment was conducted to determine whether axon terminals containing lecuine5-enkephalin target neurons in the locus coeruleus that project to the frontal cortex and, if so, what are their morphological characteristics. By light microscopy, enkephalin-labelled processes overlapped FluoroGold retrogradely labelled neurons in the locus coeruleus. By electron microscopy, retrogradely labelled perikarya and dendrites were commonly enveloped by astrocytic processes and received few afferents in the plane of section examined. However, at sites unoccupied by glial processes, abundant afferent input could be identified. In addition, some FluoroGold-labelled perikarya and dendrites lacked this glial ensheathment but were more frequently apposed by axon terminals. Of 163 FluoroGold-labelled perikarya and dendrites examined where enkephalin immunoreactivity was present in the neuropil, 42% were contacted by enkephalin-immunoreactive axon terminals. The peroxidase-labelled enkephalin terminals as well as the unlabelled terminals often contained both small, clear and large dense core vesicles. Both labelled and unlabelled terminals also formed primary symmetric synapses characteristic of inhibitory transmitters with retrogradely labelled perikarya and proximal dendrites. At times, more than one enkephalin-labelled terminal was found to converge on a common retrogradely labelled perikarya or dendrite. These results demonstrate cellular sites where enkephalin-containing afferents may directly modulate and most likely inhibit the activity of cortically projecting neurons in the locus coeruleus.

Animals↗
PubMed1996

Ultrastructural localization of calbindin-D28k and GABA in the matrix compartment of the rat caudate-putamen nuclei.

The calcium binding protein, Calbindin-D28k, is known to be localized within spiny neurons of the matrix of the dorsal striatum, caudate-putamen nuclei. This compartment is also known to contain an abundance of GABAergic neurons and to receive extensive input from excitatory limbic and cortical afferents whose activation produces rapid influxes of calcium in neuronal targets. We used electron microscopic immunocytochemistry to examine a potential role for calbindin in GABAergic neurons in the caudate-putamen nuclei. Sections of striatal tissue from acrolein-fixed adult rat brains were dual-labeled using immunoperoxidase for the localization of rabbit anti-calbindin and immunogold-silver for the localization of rat anti-GABA antibodies. Calbindin-D28k and GABA were mainly co-localized in somata and large dendrites. The peroxidase reaction product for calbindin was diffusely distributed throughout the neuronal cytoplasm, but appeared more densely localized along asymmetric, excitatory-type, postsynaptic junctions of dendritic spines, as well as saccules of smooth endoplasmic reticulum near dendritic appositions. In contrast, the immunogold-silver labeling for GABA was largely restricted to perikarya and large dendrites. Axon terminals forming symmetric junctions were also sometimes dual-labeled for calbindin and GABA. However, the majority of the calbindin-immunoreactive terminals did not contain GABA and many formed asymmetric excitatory-type synapses with either unlabeled or calbindin-labeled dendritic spines. These results suggest that, in the striatal matrix, Calbindin-D28k contributes to the immobilization of calcium (i) in selectively activated postsynaptic spines of GABAergic and possibly non-GABAergic neurons and (ii) in terminals containing GABA as well as other excitatory and inhibitory transmitters. The extent to which calbindin is able to restrict the cytosolic increases in calcium to selective sites of utilization in these neurons may have important consequences for normal synaptic function and for neuroprotection against excitoxicity.

Animals↗
PubMed1995

Morphologically heterogeneous met-enkephalin terminals form synapses with tyrosine hydroxylase-containing dendrites in the rat nucleus locus coeruleus.

Physiological and anatomical studies have suggested that the endogenous opioid peptide, methionine-enkephalin (ENK), may directly modulate noradrenergic neurons. Additionally, chronic opiate administration has been shown to increase the levels of a number of G-proteins and phosphoproteins including the catecholamine synthesizing enzyme, tyrosine hydroxylase (TH). We combined immunogold-silver localization of tyrosine hydroxylase and immunoperoxidase labeling for ENK in single sections through the nucleus locus coeruleus (LC) in the rostral pons to determine potential substrates for the divergent actions of this opioid peptide. Light microscopic analysis of ENK immunoreactivity in the LC area indicated that ENK fibers are dense and highly varicose. In coronal sections, ENK-immunoreactive processes were punctate and appeared to envelop LC-cell bodies. More rostrally, in the region of catecholamine-immunoreactive extranuclear dendrites, ENK-immunoreactive varicose processes were interdigitated with TH-labeled processes. Electron microscopy of this rostral region revealed that ENK-immunoreactive axon terminals contained small clear as well as large dense core vesicles. The large dense core vesicles (1-10/terminal) were consistently the most immunoreactive and were identified toward the periphery of the axon terminal distal to the active zone of the synapse. Unlabeled axon terminals and glial processes were the most commonly observed elements located adjacent to the plasmalemma of axons containing the labeled dense core vesicles. Axon terminals containing ENK immunoreactivity varied in size (0.3 micron to 2.0 microns) as well as formation of synaptic specializations (i.e., asymmetric versus symmetric). The ENK-labeled terminals formed synapses with dendrites with and without detectable TH immunoreactivity. These results provide the first direct ultrastructural evidence that morphologically heterogeneous terminals containing ENK immunoreactivity form synapses with catecholamine dendrites within the LC. The formation of asymmetric and symmetric synaptic specializations suggests that the opioid peptide, ENK, may be colocalized with other neurotransmitters. Furthermore, the distribution of ENK immunoreactivity in axon terminals apposed to other unlabeled afferents or astrocytic processes suggests that actions of ENK may also include presynaptic modulation of other transmitters and/or effects on astrocytes.

Animals↗
PubMed1995

Amygdala efferents form inhibitory-type synapses with a subpopulation of catecholaminergic neurons in the rat Nucleus tractus solitarius.

The central nucleus of the amygdala (CNA) integrates visceral responses to stress partially through efferent projections to portions of the medial nuclei of the solitary tracts (mNTS) containing catecholaminergic neurons. To determine anatomical sites for CNA modulation of these neurons, immunoperoxidase detection of anterogradely transported Phaseolus vulgaris-leucoagglutinin (PHA-L) or biotinylated dextran amine (BDA) was combined with immunogold-silver labeling of the catecholamine-synthesizing enzyme, tyrosine hydroxylase, in adult rat mNTS. From 350 anterogradely labeled terminals identified within the intermediate mNTS, 30% formed symmetric, inhibitory-type synapses and the remainder lacked recognized junctions as seen within a single plane of section. Of the terminals forming symmetric synapses, 16% were presynaptic to tyrosine hydroxylase immunoreactive dendrites and the remainder to unlabeled dendrites. The level of tyrosine hydroxylase immunoreactivity as assessed by density of gold-silver particles was significantly lower in dendrites receiving synaptic input from CNA efferents as compared with dendrites of the same sizes (2.0 microns 2 in mean area) which received synapses from unlabeled terminals or lacked recognizable synaptic inputs. When separately examined without regard to afferent input, the medium- and larger-sized dendrites having mean cross-sectional areas of 1-3 microns 2 also contained significantly less tyrosine hydroxylase immunoreactivity than small (< 1 micron 2) dendrites. These results suggest that CNA efferents to the mNTS inhibit non-catecholamine-containing neurons and a subpopulation of catecholaminergic neurons distinguished by their low levels of tyrosine hydroxylase. The findings also indicate that small, presumably more distal, dendrites in the intermediate mNTS may synthesize and/or release catecholamines.

Afferent Pathways↗
PubMed1995

Ultrastructural localization of delta-opioid receptor and Met5-enkephalin immunoreactivity in rat insular cortex.

The insular cortex has been implicated in the reinforcing properties of opiates as well as in the integration of responses to sensory-motor stimulation. Moreover, the delta-opioid receptor (DOR) and the endogenous opioid ligand, Met5-enkephalin (ENK) are known to be prominently distributed in insular limbic cortex. To examine the anatomical sites for opioid activation of DOR in rat insular cortex, we used immunoperoxidase for detection of an antiserum raised against a peptide sequence unique to the DOR alone, and in combination with immunogold-silver labeling for ENK. Light microscopy showed intense DOR-like immunoreactivity (DOR-LI) in pyramidal cells and interneurons in deep laminae, and in varicose processes in both superficial and deep layers of the insular cortex. Ultrastructural analysis of layers V and VI in insular cortex showed that the most prominent immunoperoxidase labeling for DOR was in dendrites. This labeling was associated with asymmetric excitatory-type junctions postsynaptic to unlabeled terminals. Dendritic DOR-LI was also distributed along selective portions of non-synaptic plasma membranes and subsurface organelles. In dually labeled sections, dendrites containing DOR-LI sometimes received synaptic input from ENK-labeled terminals or more infrequently colocalized with ENK. Other axon terminals were exclusively immunolabeled for DOR or more rarely contained both DOR and ENK immunoreactivity. Within labeled axon terminals, distinct segments of the plasma membrane and membranes of immediately adjacent synaptic vesicles showed the largest accumulation of the peroxidase reaction product for DOR. These results indicate that in rat insular cortex DOR is primarily heteroreceptive, but also serves an autoreceptive function on certain ENK-containing neurons. Our results also provide the first ultrastructural evidence that in rat insular cortex endogenous opioids interact through the DOR (1) to modulate the postsynaptic responses to other excitatory afferents and (2) to presynaptically regulate the release of other neurotransmitters. The modulatory actions on both ENK-containing and non-ENK-containing neurons may contribute significantly to the reinforcing properties of exogenous opiates acting on the DOR in limbic cortex.

Animals↗
PubMed1995

Comparative ultrastructural localization of the NMDAR1 glutamate receptor in the rat basolateral amygdala and bed nucleus of the stria terminalis.

The N-methyl-D-aspartate (NMDA)-type glutamate receptor in the basolateral amygdala (BLA) has been implicated in activity-dependent plasticity important for cortically evoked acquisition of fear-potentiated startle response. We examined the ultrastructural immunoperoxidase labeling of the R1 subunit of the NMDA receptor in the BLA of adult rats to determine the potential cellular and subcellular sites mediating the effects generated by NMDA activation. The localization was compared with that seen in the bed nucleus of the stria terminalis (BNST), the major efferent pathway from the central nucleus of the amygdala, which has a more pronounced involvement in autonomic function. Electron microscopy established that in the BLA, 68.4% (n = 177) of the profiles showing NMDAR1-like immunoreactivity (NMDAR1-LI) were dendrites, and 19.8% were distal tips of astrocytic processes. In contrast, profiles containing NMDAR1-LI (n = 262) in the BNST were more equally distributed between dendrites (37.4%) and axons (38.2%). The subcellular localization of NMDAR1 immunoreactivity was, however, similar in both regions. Our findings provide the first ultrastructural evidence that glutamate may prominently act through NMDAR1 receptors to elicit postsynaptic actions on intrinsic neurons in the BLA and BNST. The results also indicate that, in the BLA, the NMDAR1 receptor plays an important role in astrocytic function, whereas the receptor is more preferentially a presynaptic modulator in axons which terminate in or pass through the BNST.

Amygdala↗