FOS and FOSB expression in the medial preoptic nucleus pars compacta of maternally active C57BL/6J and DBA/2J mice.

C57BL/6J and DBA/2J inbred mice differ in aspects of maternal behavior and in the morphology of the medial preoptic nucleus (MPO), suggesting a possible association. DBA/2J mice have a compact subnucleus in the MPO, the MPOpc, that is sexually dimorphic and absent in C57BL/6J mice. To determine whether MPOpc cells are activated by maternal behavior, FOS and FOSB immunohistochemistry was performed on brain sections of C57BL/6J and DBA/2J mothers following the return of their pups after a separation of 2 days. In both light and dark phases of the daily cycle, stimulation of DBA/2J mothers evoked an increase in FOS- and FOSB-immunoreactivity in the MPOpc. Stimulated C57BL/6J mice, which lack the MPOpc, did not show an increase in cellular activity in the corresponding MPO region. Cells immediately lateral to the MPOpc were activated by pup stimulation, in both strains. These results suggest that MPOpc cells are active during maternal behavior, and that strain differences in maternal behavior are related to anatomical differences in the MPO.

Strain and sex differences in the morphology of the medial preoptic nucleus of mice.

The medial preoptic nucleus (MPO), which is involved in sexual and maternal behaviors, contains neuronal clusters that have been described as being sexually dimorphic in size and neuropeptide content in a variety of species. A subnucleus in DBA/2J (D2) inbred mice, called the pars compacta of the MPO (MPOpc), is absent in C57BL/6J (B6) inbred mice (Robinson et al. [1985] J. Neurogenet. 2:381-388). We report here on experiments that further characterize strain and sex differences in medial preoptic morphology in D2 and B6 inbred mice. A prominent MPOpc, located within the caudal part of the MPO and dorsal to the suprachiasmatic nucleus, was present in both male and female D2 animals but was absent from B6 animals. MPOpc neurons were darkly stained for Nissl substance and larger than neurons in the surrounding MPO. In D2 brains, galanin-immunoreactive (-ir), oxytocin-ir, vasopressin-ir, and NADPH diaphorase-positive neurons were concentrated within the MPOpc. Fewer similar neurons in the comparable region of the MPO of B6 mice suggests that the absence of the MPOpc is due to absence of these neurons rather than a less compact organization. In D2 animals, the density of galanin-ir neurons in the MPOpc was sexually dimorphic, with higher numbers of galanin-ir neurons in females. Strain differences in galanin-ir, oxytocin-ir, vasopressin-ir, and NADPH diaphorase staining appeared to be limited to the MPOpc. Cholecystokinin-immunoreactive neurons, which have been reported to be numerous in the sexually dimorphic central subdivision of the MPO of rats, were sparse in the MPO of D2 and B6 mice. Confirmation of the MPOpc as an accessory magnocellular neurosecretory nucleus was obtained by finding labeling of MPOpc neurons after injection of DiI into the posterior pituitary.

Maternal behavior in female C57BL/6J and DBA/2J inbred mice.

Inbred strains of mice exhibit different patterns of maternal behavior, providing material for studies of genetic influences on the expression of maternal behavior. Beginning 1 day after birth, maternal behavior was recorded daily for 14 days in the first and second litters of C57BL/6J (B6) and DBA/2J (D2) mothers. D2 mice had higher pup survival than B6 mice, and pup survival was higher in both strains in second litters than in first litters. D2 mothers spent more time engaged in maternal behavior, especially resting with, crouching over, and nursing pups than B6 mothers with first litters, but not with second litters. Not all measures of maternal behavior were correlated with pup survival; with both litters, B6 mothers retrieved pups faster than D2 mothers.

Development of bombesin-like and histamine-like innervation in the bullfrog (Rana catesbeiana) central nervous system.

Amphibians rely exclusively on behavioral thermoregulation to maintain body temperature within species- and developmental stage-specific critical limits. Several members of the bombesin family of peptides and histamine are included in a class of neurochemicals that have potent thermoregulatory effects in ectothermic and endothermic vertebrate species and may be involved in behavioral thermoregulation in amphibians. Because amphibians respond to environmental temperature cues differently in larval versus adult animals, we used immunocytochemistry to study developmental changes in bombesin-like (BN) and histamine-like (HA) innervation in the bullfrog brain and spinal cord. Neurons and fibers that were BN-immunoreactive and HA-immunoreactive were present in the earliest stage tadpoles examined (Gosner stage 29); BN-immunoreactive perikarya were found only in the preoptic area, posterior thalamic nucleus and in the rostroventral tegmentum of the mesencephalon. In the preoptic area, dramatic changes were observed in the number and staining intensity of BN-ir somata; neuronal labelling was greatest in tadpoles undergoing tail resorption (i.e. metamorphic climax) and was nearly absent in adults. Neurons immunoreactive to BN in the ventral mesencephalon also were developmental stage-dependent; limb-bud growth stage tadpoles had the largest numbers of labelled neurons, whereas in the adults, labelled cells were rarely visible in this area. The highest density of fibers was in the medial septum, lateral amygdala, and the optic tectum. Fewer fibers were observed within the dorsal and ventral hypothalamus, the pineal gland, and all the thalamic nuclei. Perikarya immunoreactive to HA were localized in the dorsal infundibular nucleus of the hypothalamus. Immunoreactivity was present in all developmental stages examined, and the numbers of labelled cells increased throughout metamorphosis to a maximum in adult brains. Fibers were found in the medial septum, medial amygdala, preoptic area, thalamus, pineal gland, hypothalamus and optic tectum. These results show that BN- and HA-immunoreactivities are established early in larval development, but their phenotypes are differentially expressed during larval and adult growth stages. This pattern suggests that reorganization of BN-like and HA-like neural circuitry may occur during metamorphosis and may be involved in the reported developmental changes in amphibian thermoregulation. In addition, BN-like peptides and HA may modulate other related mechanisms of amphibian thermoregulation and behaviour, such as thermal acclimation, circadian shifts in temperature selection and feeding. To what extent they are involved in amphibian thermoregulation remains to be investigated.

Effects of bombesin on behavioral thermoregulation in the bullfrog.

Bombesin is a member of a class of neuroactive chemicals that have potent thermoregulatory effects in ectothermic and endothermic vertebrate species. Bombesin-like peptides are found in the brains of ectothermic and endothermic vertebrates and have been implicated in the central nervous system modulation of behavioral thermoregulation. Amphibians rely on behavioral thermoregulation to maintain their body temperature within developmental stage-dependent critical limits. To investigate the influence of bombesin on behavioral thermoregulation, we examined the effects of central injections of bombesin on thermal habitat selection at different stages of bullfrog development. Tadpoles and adult male and female frogs were allowed to select a preferred temperature, within an aquatic thermal gradient, before and after receiving an intracerebroventricular injection of bombesin. In larval and adult female bullfrogs, bombesin administration caused a decrease in preferred temperature values. This effect was clearly dose-dependent in tadpoles. Bombesin effects were variable in adult males, probably due to an overriding stress response to handling exhibited by males. The bombesin-induced hypothermia was blocked by [D-Phe6, Des-Met14]-bombesin (6-14), ethyl amide, a bombesin/gastrin-releasing peptide receptor antagonist. These data suggest that bombesin/gastrin-releasing peptide receptors are functional in the central nervous system of larval and adult amphibians and that receptor binding can modulate thermoregulation. They raise the question: under what natural conditions is endogenous bombesin/gastrin-releasing peptide released in the brain to activate thermoregulatory behavior?

Development of arginine vasotocin innervation in two species of anuran amphibian: Rana catesbeiana and Rana sylvatica.

Arginine vasotocin (AVT) is a neurotransmitter in the amphibian central nervous system and is released from the neurohypophysis in the regulation of hydromineral balance and other homeostatic functions. Many amphibians experience drastic changes in habitat with respect to water availability during their transformation from aquatic larvae to terrestrial adults. To examine whether metamorphosis is accompanied by a reorganization of central vasotocinergic neurons, the developmental organization of vasotocin neurons and nerve fibers was studied with immunocytochemistry in the brains of bullfrogs (Rana catesbeiana) and woodfrogs (R. sylvatica). In bullfrogs, early limb-bud-stage tadpoles had AVT-immunoreactive neurons and nerve fibers in the lateral septal nucleus, amygdala, preoptic hypothalamus, suprachiasmatic nucleus, and posterodorsal tegmentum. Woodfrog larvae showed similar patterns of hypothalamic AVT immunoreactivity, although neuronal staining in the amygdala did not appear until metamorphic climax, and never appeared in septal neurons or in the posterodorsal tegmentum. Whereas the highly terrestrial R. sylvatica adults must adapt to an adult habitat with prolonged periods of dehydration, R. catesbeiana adults remain semiaquatic and, as such, need not develop extreme mechanisms for water retention. Nonetheless, vasotocinergic pathways showed developmental similarities in the two species. The early appearance of AVT innervation in both Rana suggests that AVT has neuroregulatory functions well before metamorphosis.

Interleukin-1 beta has excitatory effects on neurons of the bed nucleus of the stria terminalis.

Arginine vasopressin (AVP) is an endogenous antipyretic which acts in the ventral septal area (VSA) of the brain following its release from terminals of neurons from the bed nucleus of the stria terminalis (BST). The neurochemical mechanisms involved in the activation of these BST neurons are unknown. The present study was conducted to determine whether a naturally occurring brain cytokine (interleukin-1 beta, IL-1) selectively activates the population of BST neurons projecting to the VSA or another locus known to receive vasopressinergic input from the BST, the habenular nuclei (HAB). Single unit extracellular recordings were made from identified BST neurons in urethane-anesthetized rats. Human recombinant IL-1 applied iontophoretically or by micropressure, evoked marked excitations of long duration in 24% of all BST cells observed (n = 102 cells). Iontophoresis of sodium salicylate attenuated or reversed the effects of the cytokine in all cases tested. The selective and long-lasting excitatory actions of IL-1 on BST neurons are consistent with a direct CNS function for this cytokine. In addition, these results are compatible with a role for IL-1 in evoking AVP release from BST neurons during fever.

The effectiveness of arginine vasopressin and sodium salicylate as antipyretics in the Brattleboro rat.

The infusion of either 30 micrograms/microliters (approx. 100 micrograms/kg/h) of sodium salicylate or 10 ng/microliters (10(-5) M) arginine vasopressin within the ventral septal area of the Brattleboro rat brain reduced a centrally induced prostaglandin E1 (PGE1) hyperthermia when compared with infusions of artificial cerebrospinal fluid. Conversely, the infusion of a related peptide, oxytocin (10 ng/microliters (10(-5) M), or 33 ng/kg/h) failed to alter the rise in core temperature following the PGE1 injection. These results suggest that the vasopressin receptors reported to be present in the Brattleboro rat may respond normally to exogenously administered vasopressin, thus allowing for the antipyretic action. Moreover, the antipyretic effects of sodium salicylate suggest that aspirin-like drugs may induce the release of alpha-melanocyte-stimulating hormone which, in turn, attenuates the PGE1-evoked fever. Given recent evidence, however, which suggests that the Brattleboro rat may contain vasopressin both peripherally and within the brain, the antipyretic action of sodium salicylate may be alternatively explained through the endogenous release of vasopressin.

Single-unit activity in the bed nucleus of the stria terminalis during fever.

Arginine vasopressin, released from nerve terminals in the septal region, probably exerts endogenous antipyretic activity. A major source of vasopressin to this area is the bed nucleus of the stria terminalis (BST). In order to characterize electrophysiologically the BST-septal pathway and its potential role in the control of fever, single-unit, extracellular recordings were made from neurons in the BST of anesthetized rats. Afferent and efferent connections were identified by electrical stimulation of the medial amygdaloid nucleus and the ventral septal area (VSA). BST neurons received both inhibitory and excitatory synaptic input from the amygdala and VSA. Efferents to the VSA were identified by stimulus-evoked antidromic spike invasion. Some BST neurons were responsive to peripheral skin temperature (thermoresponsive). The activity of putative vasopressin neurons was studied during prostaglandin E1-induced fever. Although a majority of BST units was unaffected by fever, a proportion of the cells examined increased their firing rates in accordance with reported release of vasopressin in the VSA during fever.

Morphological and electrophysiological properties of a novel in vitro preparation: the electrosensory lateral line lobe brain slice.

An in vitro brain slice preparation of the electrosensory lateral line lobe (ELL) of weakly electric fish was developed. The morphology of this slice was studied and revealed that most ELL neurons and synapses retained their normal appearance for at least 10 h in vitro. The electrophysiological characteristics of the main ELL output neurons, the pyramidal cells, were measured. Extracellular electrode recordings demonstrated that pyramidal cells are capable of spontaneous, rhythmic spike activity. Intracellular recordings showed that intrinsic oscillations in membrane potential underlie the bursting behavior. The majority of pyramidal cells respond to depolarizing current pulses with an initial lag in spike firing followed by a non-accommodating, higher frequency spike train. Time and voltage-dependent properties of pyramidal cell responsiveness, as well as the effects of pharmacological blocking agents indicated that rhythmic activity and repetitive firing are dominated by a persistent, subthreshold sodium conductance (gNa) which activates at depolarizing levels and is the driving force behind the membrane potential oscillations and the sustained (non-accommodating) spike firing. In addition, a transient, outward potassium conductance (gA) is responsible for the lag in spike firing by acting as a 'brake' during the initial 50-200 ms of a depolarizing stimulus. Calcium currents and calcium-dependent potassium conductance add to the interval between spontaneous bursts but appear insufficient for spike frequency accommodation. The in vitro behaviour of pyramidal cells differs substantially from the behaviour of the same cell type in vivo. These observations raise possibilities that intrinsic membrane properties together with local synaptic interactions may regulate pyramidal cell responsiveness.

Ultrastructural studies of physiologically identified electrosensory afferent synapses in the gymnotiform fish, Eigenmannia.

Eigenmannia is a weakly electric fish that emits a constant-frequency electric organ discharge (EOD). Probability coder (P unit) and phase coder (T unit) electroreceptive afferents differentially encode changes in EOD amplitude and phase, respectively. physiologically identified T and P units were intracellularly labelled with HRP and their terminals were examined with electron microscopy to determine their postsynaptic targets. This technique reveals that phase and amplitude are relayed to first-order electrosensory neurons by two parallel but not independent pathways. P-type afferents terminate on granular interneurons, basilar pyramidals, and polymorphic cells, electrosensory lateral line lobe targets that monitor amplitude modulations, but P-type afferents do not contact spherical cells. T-type afferents relay phase information to spherical cells and thus form a separate afferent pathway. T unit terminals do not synapse directly on basilar pyramidal cells. Collateral branches from T-type afferents, however, were also found to terminate on granule and polymorphic cells, thereby adding phase information into the amplitude channel. P- and T-type afferents exhibit cellular specificity by forming synaptic junctions with different subsets of post synaptic targets in the deep neuropil. The afferent terminals make either asymmetric chemical or gap junction synapses depending on the identity of the post synaptic target. T units contacting granule cells or polymorphic cells had not been previously described. Two possible roles of adding phase to amplitude information are discussed in terms of electrolocation.

The effect of nerve activity on the distribution of synaptic vesicles.

Certain gymnotid fish (apteronotids) continuously emit a high-frequency electric-organ discharge and thus continuously drive their electroreceptor afferents at high rates. Electroreceptor afferents terminate in one lamina of the electrosensory lateral line lobe (ELL) and can be readily sampled. Normally these terminals have many small vesicles clustered adjacent to the presynaptic membrane. When afferent activity is blocked for 24 hr by an injection of tetrodotoxin (TTX) into the electroreceptor nerve, the density of vesicles adjacent to the synaptic membrane declines; the volume of the remaining vesicles increases. If the nerve of a TTX-treated fish is stimulated proximal to the injection site, these changes can be reversed. These results imply that the migration of vesicles toward the presynaptic membrane is influenced by the level of activity in the nerve.

The distribution of acetylcholinesterase and choline acetyl transferase in the cerebellum and posterior lateral line lobe of weakly electric fish (Gymnotidae).

Cholinesterase was demonstrated in the caudal lobe of the cerebellum but not the corpus cerebelli of weakly electric gymnotid fish. It had a patchy distribution in the granule cell layer and was very dense in the molecular layer; the cholinesterase staining was also dense in the contiguous molecular layer of the subjacent electrosensory region. Choline acetyltransferase was also found in far greater amounts within the electrosensory region and caudal lobe of the cerebellum than within the corpus cerebelli itself.