S.R. Vincent

GABA Neuron Systems in Hypothalamus and the Pituitary Gland

The distribution of gamma-aminobutyric acid (GABA) nerve fibers and cell bodies in the rat hypothalamus and pituitary gland was immunohistochemically examined using antibodies against the GABA-synthesizing enzyme glutamate decarboxylase (GAD). The dense network of GAD-positive nerve fibers was observed to be essentially evenly distributed throughout the hypothalamus. A plexus of GABA terminals was also demonstrated both in the median eminence and with in the posterior and intermediate lobes of the pituitary. Three distinct clusters of magnocellular GABA neurons were discovered in the posterior hypothalamus. In the addition, GAD immunoreactive cell bodies were observed in many other hypothalamic nuclei, such as the arcuate nucleus and in the perifornical region. The results provide a morphological basis by which GABA of hypothalamic origin may regulate the neuroendocrine system.

Dynorphin-immunoreactive neurons in the central nervous system of the rat

Abstract An antiserum specific for the C-terminal region of dynorphin 1–17 (DYN) was used to examine the distribution of this endogenous opioid peptide in the rat brain with the indirect immunofluorescence technique. DYN-positive nerve cell bodies and fibers were found in many nuclei in the spinal cord, medulla, mesencephalon, hypothalamus and forebrain. These findings indicate that a widespread system of DYN neurons is present in the brain distinct from the previously described enkephalin and endorphin systems.

Coexistence of somatostatin- and avian pancreatic polypeptide (APP)-like immunoreactivity in some forebrain neurons

The indirect immunofluorescence technique was used to demonstrate the coexistence of somatostatin together with avian pancreatic polypeptide-like immunoreactivity within certain neurons of the rat forebrain. Numerous neurons containing these peptides were observed in the neocortex, hippocampus, olfactory tubercle, striatum, nucleus accumbens and lateral septum. In studies of serial sections stained alternately for these two peptides, and in restaining experiments, It could be determined that in many neurons in these areas these two peptides coexisted. In other brain areas such as the anterior periventricular hypothalamus, somatostatin cells were never found to contain avian pancreatic polypeptide-like immunoreactivity. Also, within the pancreas these two peptides were never found to coexist in the same cells. The findings represent a further example of the coexistence of more than one neuropeptide within a single neuron.

Coexistence of cholecystokinin- and substance P-like peptides in neurons of the dorsal root ganglia of the rat

Using the indirect immunohistochemical technique with antisera to cholecystokinin and to substance P, the spinal dorsal horn and dorsal root ganglia of normal and colchicine-treated rats were studied. In the spinal cord a similar distribution of substance P- and cholecystokinin-positive networks in the superficial layers of the dorsal horn was observed. In the dorsal root ganglia several cholecystokinin and substance P immunoreactive cell bodies were seen in colchicine-treated rats. After elution and restaining for substance P, of sections previously stained for cholecystokinin, it was found that all cholecystokinin-positive cells also contained substance P-like immunoreactivity and vice versa.

Dynorphin-immunoreactive neurons in the autonomic nervous system

The distribution of dynorphin-like immunoreactivity in the autonomic nervous system of the rat and guinea-pig was investigated using an antiserum raised against dynorphin-(1-17). Dynorphin-like immunoreactivity was observed in fiber networks in the prevertebral sympathetic ganglia, in fibers in the enteric plexa , circular muscle layer and a few in the lamina muscularis of the mucosa of the gastrointestinal tract. After colchicine treatment dynorphin-immunoreactive cell bodies were observed both in the myenteric and submucous ganglia of the gut. The paravertebral ganglia contained occasional dynorphin-positive fibers and a few immunoreactive small intensely fluorescent cells. The distribution of the dynorphin-like immunoreactivity was compared to the distribution of enkephalin-like immunoreactivity, and it was found that in some areas there were similarities in the distribution patterns, although in other areas there were clear differences, indicating a non-identity of these two systems.

Immunohistochemical Studies of the GABA System in the Pancreas

gamma-Amino butyric acid (GABA) is a major inhibitory neurotransmitter in the mammalian brain. In the present study the indirect immunofluorescence technique was employed to localize the GABA-synthesizing enzyme glutamate decarboxylase, and the GABA-metabolizing enzyme GABA-transaminase within the rat pancreas. Both enzymes were found to occur only in the beta-cells of the islets of Langerhans. The other endocrine cell types, the exocrine tissue and the nervous elements in the pancreas did not contain either enzyme. Animals treated with the beta-cell toxins streptozotocin or alloxan showed a loss of immunoreactive cells in the islets. The results provide morphological evidence of the coexistence of GABA and insulin in the beta-cells of the endocrine pancreas.

Effects of methylene blue and LY83583 on neuronal nitric oxide synthase and NADPH-diaphorase

Methylene blue and 6-anilino-5,8-quinolinedione (LY83583) have often been used as selective inhibitors of soluble guanylyl cyclase. We report that in in vitro assays, both these compounds were potent inhibitors of rat cerebellar nitric oxide synthase activity. Methylene blue had an apparent Ki of 2.7 microM, while for LY83583 the Ki was 15.8 microM. Furthermore, methylene blue, but not LY83583, inhibited the NADPH-diaphorase histochemical reaction associated with nitric oxide synthase. Our results indicate that many of the effects of these drugs which have been attributed to inhibition of guanylyl cyclase, may derive from their direct inhibition of nitric oxide synthase activity instead.

Peripheral projections and neuropeptide coexistence in a subpopulation of fluoride-resistant acid phosphatase reactive spinal primary sensory neurons

Combined retrograde axonal tracing with the fluorescent dye Fast Blue and fluoride-resistant acid phosphatase (FRAP) histochemistry revealed that the FRAP-containing sensory neurons project to both somatic and autonomic peripheral nerves. Furthermore, the combination of indirect immunohistochemistry after colchicine treatment and FRAP histochemistry showed that a population of FRAP-containing sensory neurons are also substance P, cholecystokinin or somatostatin positive.

Immunohistochemical demonstration of plasticity in GABA neurons of the adult rat dentate gyrus

The distribution of GABA fibers within the dentate gyrus was immunohistochemically examined following lesions of the entorhinal cortex in the adult rat. A major change in the pattern of the GAD immunoreactive fibers within the molecular layer, characterized by a marked increase in the density of fibers in the outer molecular layer, was observed. This change in the lamination of the dentate GABA fibers following entorhinal lesions appeared very similar to the changes which occur in acetylcholinesterase staining following entorhinal denervation of the dentate. These results provide morphological support for the sprouting of GABA fibers in the dentate gyrus in response to perforant path destruction.

Coexistence of classical transmitters and peptides in neurones

The term “coexistence” in the context of the occurrence of several putative transmitter substances in a single neurone was perhaps first used by Brownstein et al. (1974). The authors studied neurones isolated from invertebrate ganglia and some of these cells contained 5-hydroxytryptamine (5-HT), octopamine and acetylcholine. Similar types of neurones have also been investigated in other laboratories and shown to contain more than one transmitter candidate (Kerkut et al., 1967; Hanley and Cottrell, 1974; Cottrell, 1976; Osborne, 1977). However, as discussed by Osborne (1979), in no case has evidence been presented that, if two (or more) transmitter candidates coexist in an invertebrate neurone, both (all of them), in fact, fulfil a transmitter role in that particular neurone. A different type of coexistence situation has been observed in developing autonomic neurones. Thus, such neurones may, at least under in vitro conditions, switch from a cholinergic to an adrenergic transmitter and may during a certain phase contain both acetylcholine and noradrenaline. Whether or not such a coexistence also occur in adults will still have to be established, but the reader is reminded of early views held by Burn and Rand (1965) on the occurrence of acetylcholine in noradrenergic neurones and the control of noradrenaline release by acetylcholine. For an extensive treatment of this field the reader is referred to some recent review articles by Patterson (1978) and Potter et al., (1981).