Luteinizing Hormone-Releasing Hormone Neurons which are R-etrogradely Labeled After Peripheral Fluoro-Gold Administration in the Male Ferret

This study identified luteinizing hormone-releasing hormone (LHRH)-producing neurons which have access to fenestrated capillaries in prepubertal male European ferrets. Fluoro-Gold was injected intraperitoneally to retrogradely label neurons with terminals outside the blood-brain barrier. LHRH neurons were identified by immunofluorescence using a secondary antibody tagged with tetramethylrhodamine isothiocyanate. Cell bodies which demonstrated both tetramethylrhodamine isothiocyanate and Fluoro-Gold fluorescence were defined as LHRH-producing neurons with axon terminals in regions containing fenestrated capillaries. The total number and neuroanatomical distribution of immunopositive (LHRH +) cells concurred with previous studies in the ferret in which cell bodies were diffusely distributed from rostral forebrain through caudal diencephalon, with approximately 70% of the LHRH + cell bodies located in retrochiasmatic hypothalamus. In the present study, an average of 59.8% of all LHRH+ neuronal perikarya also contained Fluoro-Gold. The majority of Fluoro-Gold filled LHRH+ neurons demonstrated only faint to moderate amounts of Fluoro-Gold when compared to other Fluoro-Gold filled neurosecretory neurons. This limited uptake of Fluoro-Gold may be due to a relative inactivity of LHRH neurons projecting outside the blood-brain barrier. Double-labeled LHRH + neurons were dispersed throughout the entire population of LHRH+ cell bodies and no apparent nuclear groups of double-labeled neurons were found. This observation suggests that the LHRH+ neurons responsible for neurosecretion into the median eminence coexist with the LHRH+ neurons responsible for intracerebral neurotransmission or neuromodulation. One distinguishable population of LHRH + neurons was consistently observed in all the brains. Only 26% of total LHRH+ perikarya within the caudal arcuate nucleus contained Fluoro-Gold, while at least 50% of LHRH+ neurons in other structures, including the rostral arcuate nucleus, contained Fluoro-Gold. Thus, in the prepubertal male ferret, the majority of LHRH cell bodies located in the caudal arcuate nucleus may be differentially regulated and/or involved in non-neuroendocrine functions.     

Central distribution of subdiaphragmatic vagal branches in the rat

In the rat, the subdiaphragmatic vagus nerves (SDX) have five major branches--the right gastric, the left gastric, the coeliac, the accessory coeliac, and the hepatic. Although these branches innervate more than the organs after which they are named, some mediate specific behavioral functions. In addition to the SDX trunk, the central stump of each of these branches was incubated in horseradish peroxidase (HRP) for 6 hours in anesthetized rats. After processing the vagal ganglia, pons, medulla, and upper cervical spinal cord of each preparation, the sections were examined for both retrogradely and anterogradely transported HRP reaction product. When only one nerve had been incubated, retrogradely labeled neurons were confined primarily to the ipsilateral ganglion, medulla, and spinal cord. Within the brain, a few labeled neurons occurred within the nucleus ambiguus (NA) and the reticular formation caudal to the NA, but the vast majority appeared in the dorsal motor nucleus of the vagus (DMX). The axons of most labeled neurons in the NA distributed in the gastric branches; those from cells caudal to the NA, probably distributed in the coeliac branch. Most labeled DMX cells also distributed with the gastric branches. Those on the lateral tip of the right DMX, however, had axons in the coeliac branch; those on the left DMX tip, in the accessory coeliac. After incubation of the SDX trunk, anterograde HRP reaction product occurred in the caudomedial nucleus of the solitary tract (NST) just rostral and subjacent to the area postrema (AP). Unlike the retrograde label, anterograde reaction product was bilateral, but always weaker contralaterally. Within the SDX distribution, the afferent axons from the gastric branches exhibited one pattern of termination; those from the coeliac, accessory coeliac, and hepatic branches, another. The gastric branch distributions began dorsolaterally in the SDX termination zone and continued caudally beneath the AP. Immediately subjacent to the AP, gastric branch terminals were never dense and the entire distribution faded at the level of the obex. The coeliac and accessory coeliac distributions began dorsomedially within the SDX termination zone and intensified caudally in a thin band immediately subjacent to the AP. The densest label was associated with the caudal half of the AP, but the distribution thinned rapidly caudal to the obex. The hepatic distribution was similar to that of the coeliac branches but never achieved similar density. Physiological and behavioral data correlate with the anatomical picture in that the efferent functions appear to be more densely localized than the afferent functions.     

The GABAB antagonist phaclofen inhibits the late K-dependent IPSP in cat and rat thalamic and hippocampal neurones

Phaclofen (0.5–1 mM) reversibly inhibited the late, bicuculline resistant, K⁺ dependent IPSP recorded in projection cells of the cat and rat dorsal lateral geniculate nucleus and in rat hippocampal CA1 pyramidal neurones. At the same concentrations, phaclofen reversibly blocked the K⁺ dependent, bicuculline insensitive hyperpolarization evoked by GABA and baclofen but had no effect on the GABA_A IPSP. These results represent conclusive evidence that GABA_B receptors mediate the late K⁺ dependent IPSP in cortical and subcortical neurones.     

The pelvic innervation in the rat: different spinal origin and projections in Sprague-Dawley and Wistar rats

WGA-HRP was applied to the pelvic and pudendal nerves of Sprague-Dawley and Wistar rats to compare the segmental levels of the resulting labeling. L₆ and S₁ were the segme its at which the sacral parasympathetic nucleus and the denser primary afferents occurred in Sprague-Dawley rats. The levels found in Wistar rats were S₁ and S₂, Thus indicating a disparity between both strains of rats in the spinal level of the sacral parasympathetic nucleus and the primary afferents.     

Androgen and Estrogen Effects on Vasopressin Messenger RNA Expression in the Medial Amygdaloid Nucleus in Male and Female Rats

Vasopressin messenger RNA (AVP mRNA) expression in the medial amygdala and bed nucleus of the stria terminalis (BST) is almost completely dependent on gonadal steroids. In the BST, the effects of gonadal steroids on AVP mRNA expression are sexually dimorphic. Males have more cells that express AVP mRNA and more AVP mRNA per cell than females. Here we test whether this is also true for the MA. In gonadectomized rats that were treated with testosterone, males had more cells that were labeled for AVP mRNA than females. However, the labeling per cell did not differ between males and females. To assess contribution of testosterone metabolites to these differences, male and female rats were gonadectomized and implanted with empty tubing, or tubing filled with dihydrotestosterone (DHT), estradiol (E), or E plus DHT (E + DHT). The pattern of steroid effects on AVP mRNA expression in the MA was similar in both sexes. Hardly any labeled cells were found in rats with empty implants or rats treated with DHT. Significantly more labeled cells were found in rats treated with E, and even more cells in rats treated with E + DHT. The number of AVP mRNA-labeled cells was higher in males than in females for E as well as E + DHT treatment, but the labeling per cell did not differ between sexes. These data suggest that the number of MA cells that can express AVP mRNA is higher in males than in females, but the estrogen and androgen responsiveness of individual AVP mRNA-expressing cells in the MA does not differ between sexes.     

Involvement of the PVN and BST in 1K1C hypertension in the rat

The paraventricular nucleus of the hypothalamus (PVN) and the bed nucleus of the stria terminalis (BST) are two sources of central nervous system (CNS)-derived arginine vasopressin (AVP), a nonapeptide that has been implicated in central autonomic regulation and in particular in cardiovascular regulation, through its actions within the CNS. These experiments were designed to determine if either the PVN or the BST were involved in the development of Goldblatt one-kidney one-clip (1K1C) hypertension in the rat. In order to test this hypothesis, ibotenic acid lesions of the PVN or electrolytic lesions of the BST were undertaken in both normotensive (sham-operated) rats and in 1K1C rats. In both cases the development of 1K1C hypertension was inhibited over the 18–21 days following surgery. Lesions of the PVN did not alter normal blood pressure regulation in the sham-operated animals, whereas lesions to the BST did affect normal blood pressure regulation, resulting in a dramatic increase in blood pressure during the initial days following surgery. These studies suggest that the PVN and BST are involved in the development of 1K1C hypertension in the rat, moreover the BST may also play a role in central cardiovascular control.     

Differential regulation of oxytocin and vasopressin messenger ribonucleic acid levels by gonadal steroids in postpartum rats

We previously reported that sequential estradiol and progesterone exposure followed by progesterone withdrawal increases oxytocin (OT), but not arginine vasopressin (AVP), messenger ribonucleic acid (mRNA) in the hypothalamic paraventricular nucleus (PVN) of the rat. Substitution of testosterone for progesterone and subsequent testosterone withdrawal in the estrogen-primed rat increases PVN AVP mRNA levels. At the end of pregnancy (day 21), rats are exposed to high estrogen and declining progesterone and testosterone concentrations. Coincident with these changes in circulating gonadal steroid hormones are increases in OT and AVP mRNAs. If progesterone levels are sustained at term, OT levels are attenuated and if testosterone is sustained, AVP mRNA levels are attenuated. Immediately postpartum, however, OT and AVP mRNA levels decline compared to term levels. To further determine the role of estrogen in the regulation of OT and AVP mRNAs, we performed two experiments. In the first experiment, we administered estrogen during the peripartum period to determine if estrogen supplementation prevents the relative attenuation of OT and AVP mRNAs that is seen after parturition. Day 18 pregnant rats were given estradiol-filled or empty capsules and sacrificed on day 2 of lactation. By Northern analysis, significant differences in PVN AVP, but not OT, mRNA were found between the estrogen- and sham-treated lactational animals, P < 0.02. In the second experiment, we determined if sustaining estrogen after progesterone is removed in steroid-treated ovariectomized rats is essential for the increase in OT mRNA. Ovariectomized rats were given either empty capsules or sequential estradiol- and progesterone-filled capsules and both were sustained for 12 days. When progesterone-filled capsules were removed, estradiol-filled capsules were either removed or left in place, and the animals were sacrificed 48 h later. PVN OT mRNA was analyzed by Northern blot hybridization. OT mRNA was increased in both of the steroid-treated groups to the same degree, compared to sham-treated animals, P = 0.04. In summary, estrogen supplementation during early lactation prevents the attenuation of PVN AVP, but not OT, mRNA after parturition. In the estrogen-primed ovariectomized rat, it is not necessary to sustain estrogen to see the effects of progesterone withdrawal upon PVN OT mRNA.     

The effect of adrenalectomy on stress-induced c-fos mRNA expression in the rat brain

Previously, we determined the pattern of stress-induced c-fos mRNA expression throughout the brain in order to gain further insight into the identification of the neural circuits mediating stress-induced regulation of the hypothalamic-pituitary-adrenal axis. In the present study, we determined if rapid effects of increased glucocorticoid levels after stress contribute to changes in c-fos mRNA expression. To this end, stress-induced c-fos expression was characterized in adrenalectomized (ADX) or adrenalectomized and corticosterone replaced (ADX/B) male rats. Animals were sacrificed 30 min post-onset of a 10 min swim stress, and in situ hybridization histochemistry was used to detect c-fos mRNA throughout the brain. The pattern of c-fos induction in the ADX and ADX/B animals was similar to that observed in the sham operated animals. Additionally, densitometric measurements were made to quantify the c-fos response in the paraventricular nucleus of the hypothalamus and the CA1/2 region of the hippocampus. We found that ADX did not alter the magnitude of the c-fos response to stress in these areas, but there was a slight dampening of the response in ADX/B animals. In sum, these results suggest that the pattern of c-fos expression observed 30 min post-stress is independent of stress-induced increases in circulating glucocorticoid concentrations.     

Metabotropic Glutamate Receptors in the Rat Nucleus Accumbens

The effects of glutamate metabotropic receptors (mGluRs) on excitatory transmission in the nucleus accumbens were investigated using electrophysiological techniques in rat nucleus accumbens slices. The broad-spectrum mGluR agonist (1S,3 R )-1-aminocyclopentyl-1,3-dicarboxylate, the mGluR group 2 selective agonists (S)-4-carboxy-3-hydroxyphenylglycine, (1S,3S)-ACPD) and (2S,1'S,2'S)-2-(2'-carboxycyclopropyl)glycine (L-CCG1), and the mGluR group 3 specific agonist L-2-amino-4-phosphonobutyrate (L-AP4) all reversibly inhibited evoked excitatory synaptic responses. The specific group 1 mGluR agonist (R,S)-3,5-dihydroxyphenylglycine [(R,S)-DHPG] did not depress transmission. Dose-response curves showed that the rank order of agonist potencies was: L-CCGI > L-AP4 > (1 S,3S)-ACPD. Group 2 and 3 mGluRs inhibited transmission via a presynaptic mechanism, as they increased paired-pulse facilitation, decreased the frequency of miniature excitatory postsynaptic currents and had no effect on their amplitude. The mGluRs did not inhibit transmitter release by reducing voltage-dependent Ca²⁺ currents through N- or P-type Ca²⁺ channels, as inhibition persisted in the presence of a-conotoxin-GVIA or Aga-IVA. The depression induced by mGluRs was not affected by specific antagonists of dopamine D1, GABA-B or adenosine A1 receptors, indicating direct effects. Finally, (13,s)-DHPG specifically blocked the postsynaptic afterhyperpolarization current (I_ahp). Our results represent the first direct demonstration of functional mGluRs in the nucleus accumbens of the rat.     

Projections from the renal nerve to the cat''s lateral somatosensory thalamus

The representation of the kidney in the lateral somatosensory thalamus was mapped using electrical stimulation of the renal nerve in pentobarbitone-anesthetized cats. Ninety-five of 197 thalamic neurons studied responded to renal nerve stimulation. The responsive neurons were located in the periphery of the ventral posterolateral nucleus (42%, VPL_p) and the neighboring dorsal and lateral aspects of the posterior complex (58%; POd and POl). No visceroceptive neurons were found within VPL proper. The mean response latency of the thalamic neurons to electrical nerve stimulation was 9.5 ± 2.6 ms (mean ± S.D.), suggesting an involvement of A δ, and possibly A β fibers in the primary afferent pathway. The visceroceptive neurons were further characterized with innocuous mechanical stimulation of the body surface, and for 94 of the 95 neurons a somatic receptive field could be determined. Of these, 35% were located on the lower back and belly, i.e., the dermatomes of the lower thoracic and upper lumbar spinal projection areas of the renal nerve. 52% of the somatic receptive fields were located on the contralateral foot, thigh, tail, or hind leg (lower lumbar, sacral and coccygeal dermatomes) and 13% covered the arm and upper body (upper thoracic and lower cervical dermatomes). Comparison between the thalamic representations of the renal and pelvic nerves showed that both covered comparable areas adjacent and around, but not within VPL proper. It is concluded that VPL_p, POd and POl play a role in processing visceral, possibly including nociceptive, information from the kidney of the cat.