Nancy E. Sirett

Responses from osmosensitive neurons of the rat subfornical organ in vitro

Extracellular recordings were made in vitro from 212 single units in the rat subfornical organ (SFO) and 54 single units in the rat medial preoptic area (MPO). Units were exposed to solutions made hyper-osmotic or hypo-osmotic by 1.4-11%. A reversible 30% or greater change in frequency followed the osmotic challenge in tests of 66% of units in the SFO and 46% of units in the MPO. Responses consisted of increases in frequency (excitations) or decreases in frequency (inhibitions) and were either sustained for the whole test period or of a transitory nature. Units responded to either hyperosmotic (SFO, 19%; MPO, 43%) or to hypo-osmotic changes (SFO, 30%; MPO, 28.5%) or to both (SFO, 51%; MPO, 28.5%). The response pattern of the SFO and MPO was significantly different (chi 2 54.0, 3df, P = 0.0001). In both the SFO and MPO the stimulus to which the units responded was a change in tonicity. This was indicated by the findings that similar responses were evoked by hyperosmotic changes made with either mannitol or NaCl and there was no response to solutions containing urea, either as an additive, or as a substitute for NaCl. In the SFO, in the presence of synaptic blockade produced by raising the Mg concentration in the bathing solution to 15 mM, the frequency of 19/27 units fell significantly. Responses of 40% of units to osmotic pressure changes were blocked indicating these responses were synaptically evoked. The responses which survived synaptic blockade when compared with pre-blockade responses were more often transient (P less than 0.02) and more often inhibitions. Post blockade there were also significantly more responses in the SFO to hypo-osmotic than to hyper-osmotic changes (P = 0.01). Our results suggest that while an ability to change their firing rate in response to small changes of osmotic pressure may be a general property of neurons, the neurons of the SFO are specialised for the detection of changes in the extracellular osmotic pressure.

Localization of immunoreactive angiotensin II in the hippocampus and striatum of rat brain

Angiotensin II (ANG II) was estimated by radioimmunoassay in extracts of rat brain. In extracts of whole brain the mean content was 108 +/- 16 fmol/g and estimates of ANG II in kidney and plasma were similar to previous reports. A regional distribution of ANG II was determined. Hippocampus had the highest concentration and cortex the lowest. The concentrations relative to cortex were: hippocampus, 8; striatum, 5; cerebellum, 4; hypothalamus:thalamus:septum:midbrain (HTSM), 3; and medulla, 3.

Angiotensin binding and pressor activity in the rat ventricular system and midbrain

Abstract Specific angiotensin II (AII) binding capacity was investigated in regions of rat brain thought to be involved in the central pressor effects of AII. In the midbrain specific angiotensin II binding capacity was principally localised in the superior colliculi. When the superior colliculi was divided into rostral and caudal portions, the caudal portion had a higher binding capacity than the rostral portion. In the hypothalamus AII binding capacity was only a third of that in the superior colliculi and was localised principally in the lateral hypothalamus. The medial hypothalamus and the preoptic recess each had about a quarter of the binding capacity of the lateral hypothalamus. To investigate whether AII binding capacity could be correlated with pressor activity AII was applied locally (local) to the surface of the superior colliculi (5 μg/5 μl saline) of rats with femoral artery cannulas and anaesthetised with pentobarbitone. These responses were compared with responses to the same dose given intraventricularly (IVT). When the surface of the midbrain was intact there was a pressor response to the local application not significantly different in magnitude and latency from the response to IVT AII. The midbrain was aspirated at the site of AII application and pressor responses to AII evoked during aspiration. As the midbrain lesion was extended the latency of the local pressor response was prolonged (P

Extracellular recording in rat area postrema in vitro and the effects of cholinergic drugs, serotonin and angiotensin II

Spontaneous extracellular action potentials were recorded from rat area postrema explants in vitro for up to 6 h at 35 degrees C. Their geometric mean frequency was 4.4 +/- 1.7-11 Hz (n = 120) and they were most often recorded caudal to the obex. The frequency of spontaneously discharging units could be increased three-fold by raising the KCl concentration from 5 to 15 X 10(-3) M but a claimed non-specific excitant of neurones, L-glutamic acid at 10(-7)-10(-3) M was without effect. Carbamylcholine at 10(-9)-10(-7) M increased the frequency of spontaneous units (12/13 trials) as did 10(-7) M neostigmine sulphate (14/14 trials). The effects of carbamylcholine and neostigmine were additive and were blocked by atropine sulphate at 10(-6) M (18/18 trials). Atropine also stopped the discharge of spontaneous units while D-tubocurarine did not affect unit discharge frequency. It is suggested that units responding to cholinergic drugs have an afferent input from the dorsal vagus. A number of putative transmitters, serotonin (10(-9)-10(-7) M), angiotensin II (0.5 X 10(-10)-0.5 X 10(-9) M) and dopamine (10(-9)-10(-5) M) which on indirect grounds are thought to affect area postrema neurones, were without effect on unit discharge frequency.

LH-RH in picomole concentrations evokes excitation and inhibition of rat arcuate neurones in vitro

A medial sagittal brain slice was developed, which enabled electrophysiological recording from spontaneously active neurones adjacent to the infundibular recess of the rat arcuate nucleus. Luteinizing hormone-releasing hormone (LH-RH) (10 nM-10 pM) significantly altered the frequency of 21 out of 31 units tested, exciting 14 and inhibiting 7 others. The excitatory responses were often not maintained during the exposure to LH-RH but either returned to pre-exposure frequency or displayed an inhibition of discharge. A neural model incorporating recurrent inhibition of LH-RH-excited neurones is proposed to explain these responses, thought to be endogenously evoked by collateral innervation of the arcuate nucleus by medial preoptic neurones projecting to the median eminence.

Neurotensin excites neurons in the arcuate nucleus of the rat hypothalamus in vitro

The effects of neurotensin (NT) on 41 spontaneously active neurons in the rat hypothalamic arcuate nucleus (ARC) were determined using an in vitro brain slice technique. Addition of NT (1 microM to 10 nM) to the perifusing medium allowed the action of known concentrations of NT to be examined. Concentrations of peptide greater than 10 nM evoked excessive depolarization of sensitive neurons resulting in long periods of postexcitatory inhibition. At 10 nM, NT evoked reversible and repeatable excitatory (20%) and inhibitory (25%) responses. When synaptic activity was blocked by lowering the Ca2+ and raising the Mg2+ concentration of the medium only excitatory responses (25% of units tested) were observed. These results demonstrate a potent excitatory action of NT within the hypothalamic ARC at the single cell level.

Luteinizing hormone release in the anaesthetised cat following electrical stimulation of limbic structures

The release of luteinizing hormone (LH) in response to electrical stimulation of limbic centres, namely the medial preoptic region (MPO) medial basal hypothalamus (MBH) and the medial amygdala (AME) has been studied in the anaesthetised gonadectomized cat. Chronically gonadectomized cats were anaesthetised with pentobarbitone or Althesin and paired bipolar stimulating electrodes were aimed at the MPO, AME or MBH. The effect of electrical stimulation of these regions on the secretion of LH was studied by radio-immunoassay of LH in serial blood samples taken before, during and after stimulation. No change in plasma LH in response to electrical stimulation was ever recorded during pentobarbitone anaesthesia. During Althesin anaesthesia stimulation in the MPO more often than not resulted in a peak of LH release during stimulation. A peak release of LH during stimulation was also recorded when electrodes were placed in the arcuate-median eminence region of the MBH. The time-course of these peaks in LH secretion was similar to the time-course of the plasma LH responses recorded following a single intravenous injection of gonadotrophin-releasing hormone (GnRH). In contrast, electrodes placed in AME had no effect on plasma LH during electrical stimulation, but immediately after stopping it, a small LH peak was recorded. The time-course of these responses suggests a pulse release of GnRH, the rapid response to MPO and MBH stimulation possibly being the result of a direct action on GnRH neurons while the delayed AME response may be produced by AME projections to the GnRH release system. These responses could be likened to the surge of LH which in the cat occurs post-coitus.

Electrophysiological evidence for a projection from medial prefrontal and anterior limbic cortex toward the medial preoptic area in the cat

SummaryNeurons in cat medial prefrontal cortex, anterior limbic cortex and possibly the indusium griseum were identified by antidromic invasion as having axonal projections towards the medial preoptic region, using both macro- and microstimulation techniques. These projecting axons were found to be of slow conduction velocity (0.2–4.8 m/s) and to in some cases also send branches towards the anteromedial thalamus, mediodorsal thalamus, ventromedial tegmentum, basolateral amygdala or medial forebrain bundle. Threshold-depth curves for axons excited by microstimulation in the medial preoptic region were very steep, with proportionality constants of 0.3–7.1 μm/μA. Calculations based on the threshold-depth curves confirmed that microstimulation was most probably only activating axons within the MPO, and current spread to lateral fibers of passage following macrostimulation in the MPO was not detected in the branching studies.

Responses of cat preoptic neurons to stimulation of the medial frontal cortex and the medial basal hypothalamus

SummaryResponses of single preoptic neurons to electrical stimulation of the medial frontal cortex, the mediobasal hypothalamus (MBH) and the medial forebrain bundle (MFB) were recorded in anaesthetised cats. Single pulse stimulation of the medial frontal cortex orthodromically drove 96 otherwise quiescent preoptic neurons, which were found more frequently in the dorsal preoptic region, inhibited 53% of the spontaneously active preoptic neurons and excited 16%. Testing of cortically influenced preoptic neurons with MBH or MFB stimulation resulted in antidromic invasion of 6% (MBH) and 9% (MFB). Convergence of orthodromic inputs from medial frontal cortex and MBH was detected in 78% of spontaneously active preoptic neurons, and three-way convergence including input from MFB was noted in 17% of neurons tested with all stimulators. Some cortex-responsive neurons were found to also respond to vaginal or anal probing, paw squeezing and haemorrhage. The role of this input to the preoptic region from medial frontal cortex remains to be elucidated, but may include neuroendocrine, behavioural and homeostatic functions.

Does the precommissural fornix excite neurons in the cat dorsal septum which project to the medial preoptic region

SummaryThe possibility that effects of fornix stimulation on units in the medial preoptic region (MPO) may be via an interposed neuron in the dorsal septum was investigated using electrophysiological techniques. We found a reciprocal monosynaptic linkage between the MPO and the dorsal, medial and fimbrial septal nuclei. When recording in the MPO, stimulation of the dorsal septum affected 59% of spontaneously active units. Of all MPO units synaptically excited by stimulation of the dorsal septum 33% also had an input from the fornix. When recording in the dorsal septum during stimulation of the MPO and fornix, units synaptically driven by, or antidromically invaded following MPO stimulation, were found in caudal regions of the DS and units excited by stimulation of the fornix were located in more rostral regions. Units excited by stimulation of the fornix were never antidromically invaded following stimulation in the MPO, so there was no direct pathway for fornix excitation in the dorsal septum to reach the MPO.