Nicholas M Mellen

Nicholas M Mellen
University of Louisville | UL · Department of Pediatrics

PhD

About

29
Publications
1,684
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1,298
Citations
Additional affiliations
July 1993 - June 2004
University of California, Los Angeles
Position
  • PostDoc Position

Publications

Publications (29)
Conference Paper
Introduction: The effect of sustained hypoxia (SH) on brain metabolism has been well studied. However less is known about intermittent hypoxia (IH), a hallmark of obstructive sleep apnea (OSA), associated with increased risk for stroke, outcome severity and functional consequences. Hypothesis: Impaired glutamate homeostasis after IH may underlie i...
Article
Full-text available
Hypoxia alters cellular metabolism and although the effects of sustained hypoxia (SH) have been extensively studied, less is known about chronic intermittent hypoxia (IH), commonly associated with cardiovascular morbidity and stroke. We hypothesize that impaired glutamate homeostasis after chronic IH may underlie vulnerability to stroke-induced exc...
Article
Full-text available
Neurons that control breathing are distributed along the ventral respiratory column (VRC) in the ventrolateral brain stem, extending from the pons to the spinomedullary junction. These networks can be isolated in the en bloc brainstem-spinal cord preparation, which remains spontaneously active for hours under physiological conditions, but which onl...
Article
Full-text available
Although the mammalian locomotor CPG has been localized to the lumbar spinal cord, the functional-anatomical organization of flexor and extensor interneurons has not been characterized. Here, we tested the hypothesis that flexor and extensor interneuronal networks for walking are physically segregated in the lumbar spinal cord. For this purpose, we...
Article
Recent studies in vivo and in vitro suggest that both respiratory rhythmogenesis and its central chemosensory modulation arise from multiple, mechanistically and/or anatomically distinct networks whose outputs are similar. These observations are consistent with degeneracy, defined as the ability of structurally distinct elements to generate similar...
Article
Full-text available
Endogenous burster neurons (EBs) have been found at the level of the facial nucleus (VIIn), and 500 mum caudally, within the pre-Bötzinger complex (preBötC). They have been proposed as either causal to or playing no role in respiratory rhythmogenesis. Little is known about their broader distribution in ventrolateral medulla. Here, a Ca(2+) indicato...
Article
Bath-applied membrane-permeant Ca(2+) indicators offer access to network function with single-cell resolution. A barrier to wider and more efficient use of this technique is the difficulty of extracting fluorescence signals from the active constituents of the network under study. Here we present a method for semi-automatic region of interest (ROI)...
Article
Slice preparations isolate functional networks, permitting single unit recording under visual control, and the use of fluorescent indicators. Circuits of interest often lie at a tilt in both the rostrocaudal and ventrodorsal axis, thus exposing circuits of interest at the cut surface of a slice would require a device for tilting a preparation along...
Article
Recent studies have shown both the pFRG and the preBötC are sufficient to generate respiratory rhythm, and are hypothesized to do so via distinct mechanisms (Onimaru and Homma 2003; Mellen, Janczewski, Bocchiaro and Feldman 2003). The coexistence of mechanistically distinct, functionally matching networks (defined as degeneracy, Edelman and Gally 2...
Article
Full-text available
In mammals, respiration-modulated networks are distributed rostrocaudally in the ventrolateral quadrant of the medulla. Recent studies have established that in neonate rodents, two spatially separate networks along this column-the parafacial respiratory group (pFRG) and the pre-Bötzinger complex (preBötC)-are hypothesized to be sufficient for respi...
Article
In mammals, expiration is lengthened by mid-expiratory lung inflation (Breuer-Hering Expiratory reflex; BHE). The central pathway mediating the BHE is paucisynaptic, converging on neurones in the rostral ventrolateral medulla. An in vitro neonatal rat brainstem-lung preparation in which mid-expiratory inflation lengthens expiration was used to stud...
Article
We have analysed the action of the neuromodulatory catecholamine, dopamine (DA), on the lumbar locomotor network using an isolated in vitro newborn rat spinal cord preparation. We have also attempted to determine the respective contribution of the D1- and D2-like receptors on the dopamine-mediated effects. Bath application of DA-induced slow locomo...
Article
Current consensus holds that a single medullary network generates respiratory rhythm in mammals. Pre-Bötzinger Complex inspiratory (I) neurons, isolated in transverse slices, and preinspiratory (pre-I) neurons, found only in more intact en bloc preparations and in vivo, are each proposed as necessary for rhythm generation. Opioids slow I, but not p...
Article
Full-text available
This study was designed to examine the possibility that respiratory arrest during hypothermia occurs at the level of premotor or motor neurons rather than at the level of the central rhythm generator itself. Specifically, we sought to determine the consequences of hypothermic cooling until respiratory arrest, and subsequent rewarming, on neurons in...
Article
Full-text available
The isolated neonatal rat medulla generates respiratory-related rhythms recorded from cervical spinal cord ventral roots. When lungs and their vagal innervation are retained, respiratory activity is modulated by lung mechanoreceptor feedback: transient lung inflation triggered off inspiratory onset (phasic inflation) shortens inspiration and increa...
Article
Full-text available
The normal breathing rhythm in mammals is hypothesized to be generated by neurokinin-1 receptor (NK1R)-expressing neurons in the preBötzinger complex (preBötC), a medullary region proposed to contain the kernel of the circuits generating respiration. If this hypothesis is correct, then complete destruction of preBötC NK1R neurons should severely pe...
Article
Full-text available
Normal respiratory rhythm in mammals is hypothesized to be generated by NK1R neurons in the preBotC. Ablation of these neurons in adult rats was predicted to severely perturb normal breathing rhythm. We directly injected substance P conjugated saporin (SP-SAP) into the preBotC to selectively kill NK1R neurons [1]. Four to five days postinjection in...
Article
Full-text available
This study was designed to examine the possibility that respiratory arrest during hypothermia occurs at the level of premotor or motor neurons rather than at the level of the central rhythm generator itself. Specifically, we sought to determine the consequences of hypothermic cooling until respiratory arrest, and subsequent rewarming, on neurons in...
Article
Full-text available
The pre-Botzinger Complex is coextensive with a population of SP and DAMGO-sensitive inspiratory neurons, retained in the transverse slice preparation, which has been extensively used to study mechanisms for respiratory rhythm generation. Here, qualitatively different effects of peptidergic modulation were observed in two preparations reduced, and...
Article
Full-text available
In intact mammals, lung inflation during inspiration terminates inspiration (Breuer-Hering inspiratory reflex, BHI) and the presence of lung afferents increases respiratory frequency. To test whether these responses could be obtained in vitro, a neonate rat brain stem/spinal cord preparation retaining the lungs and their vagal innervation was used....
Article
We frame our use of nonlinear dynamic techniques historically, and show how evidence of phase synchronization between different lamprey motor roots is investigated using a mutual information measure. We also demonstrate a new approach for symbolically representing time series in terms of generic periodic points with an example.
Article
We examined intrinsic and extrinsic factors affecting phrenic motoneuron (PMN) excitability in neonatal rats. Using an in vitro brainstem-spinal cord en bloc, 127 PMNs were recorded under whole-cell patch-clamp conditions. Inspiratory synaptic drives and passive membrane properties, including whole-cell membrane capacitance (Cm), input resistance (...
Article
Respiration is modulated by lung mechanoreceptor feedback in vivo on a cycle-to-cycle basis. We replicated this modulation in vitro and tested four stimulus protocols to identify which of these most closely replicated in vivo responses to lung mechanoreceptor activation in mammals. We activated pulmonary vagal afferent pathways by electrical stimul...
Article
1. Cycle-to-cycle fluctuations in cycle periods and intersegmental burst delays of ventral root activity were studied during stable fictive swimming in the adult lamprey spinal cord. High spatial resolution was obtained by recording from 16 ventral roots, 1/2 on each side of the spinal cord. For ipsilateral ventral roots, correlations between cycle...

Questions

Question (1)
Question
details:
organotypic hippocampus-cortex slices harvested at P4, washed in Hank's balanced salt solution, then maintained for 12 days in culture medium (50% minimum essential medium, 25% heat-inactivated horse serum, 25% hank's balanced salt solution, supplemented with (in mM) glucose 36, glutamine 2, HEPES 25, and 1% antibiotic/antimicotic, adjusted to pH=7.3, and filtered), at 37 deg C and 5% CO2; medium changed every 3 days.
Dye loading: the mesh on which the tissue is cultured is cut out of the well, and transferred to a sylgard disk, where it is held in place by beads of crazy glue around the edge of the disk. The preparation is then lowered into 6 ml aCSF ( in mM: dextrose 30, CaCl2 1.2, KCl 3, NaCl 136, NaHPO4 1.25, MgCl2 1,NaHCO3 10) with 20 uL of 100 mM fluo 8 L-AM in a solution of 15% pluronic DMSO, aerated with a fine stream of bubbles of 95% O2, 5%CO2, for 40 min at 23 deg C.
We then move the tissue to the recording chamber, through which the following aCSF flows: in mM: NaCl 125.4, NaHCO3 26, KCl 5, KH2PO4 1.24, MgSO4 1.3, CaCl2 1.5, dextrose 10. The solution is warmed to 27 deg C and flows at 4 ml/min. 
We begin optical recording immediately after the prep is moved to the chamber. We expose the slices to 20 ms  of 450 nm wavelength light.
In some preps there is spontaneous activity,  and stimuli produce robust firing in subsets of cells. In other preps there is no spontaneous or evoked activity, but when 20 mM KCl is applied a wave of neuronal activity can be recorded optically.
So this is our problem: in slices harvested from the same animal, maintained in different wells but the same media under identical conditions, we get "normal" acitivity, or a complete lack of response, but the high K+ response shows that the tissue is viable.
We are confident that this variability is due to small differences in loading/culturing/cutting conditions.
Can anyone suggest what step in this process must be most tightly controlled to minimize this kind of variability?
Alternatively, is this variability (1 "good" slice out of 3 slices) just the way it is with organotypic cortex-hippocampal slices?

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Projects

Project (1)
Project
this will be the platform for characterizing the relationship between synaptic function and network dynamics in hippocampus. This project only makes sense is hippocampal function is not impacted by incorporation of GCaMP6F, so I'm gearing up to do behavioral studies in these mice to assess learning and memory.