Davide Dulcis

Neurotransmitter Switching in the Adult Brain Regulates Behavior

Daylight Determines Dopamine Expression of the appropriate neurotransmitters is essential for the function of neural circuits. Can neurons change their transmitter phenotype to deal with alterations in the environment? Dulcis et al. (p. 449; see the Perspective by Birren and Marder) exposed adult rats to different photoperiods mimicking summer and winter daylengths. Neurotransmitter expression switched between dopamine and somatostatin in hypothalamic neurons that regulate release of corticotropin-releasing factor. Transmitter switching occurred at the transcriptional level and was accompanied by changes in postsynaptic receptors. Switching rats from long days to short days causes neurons to change their neurotransmitters, and this changes behavior. [Also see Perspective by Birren and Marder] Neurotransmitters have been thought to be fixed throughout life, but whether sensory stimuli alter behaviorally relevant transmitter expression in the mature brain is unknown. We found that populations of interneurons in the adult rat hypothalamus switched between dopamine and somatostatin expression in response to exposure to short- and long-day photoperiods. Changes in postsynaptic dopamine receptor expression matched changes in presynaptic dopamine, whereas somatostatin receptor expression remained constant. Pharmacological blockade or ablation of these dopaminergic neurons led to anxious and depressed behavior, phenocopying performance after exposure to the long-day photoperiod. Induction of newly dopaminergic neurons through exposure to the short-day photoperiod rescued the behavioral consequences of lesions. Natural stimulation of other sensory modalities may cause changes in transmitter expression that regulate different behaviors.

Glutamatergic Innervation of the Heart Initiates Retrograde Contractions in Adult Drosophila melanogaster

The adult abdominal heart of Drosophila melanogaster receives extensive innervation from glutamatergic neurons at specific cardiac regions during metamorphosis. Here, we show that the neurons form presynaptic specializations, as indicated by the localization of synaptotagmin and active zone markers, adjacent to postsynaptic sites that have aggregates of glutamate IIA receptors. To determine the role of this innervation in cardiac function, we developed an optical technique, based on the movement of green fluorescent protein-labeled nerve terminals, to monitor heart beat in intact and semi-intact preparations. Simultaneous monitoring of adjacent cardiac chambers revealed the direction of contractions and allowed correlation with volume changes. The cardiac cycle is composed of an anterograde beat in alternation with a retrograde beat, which correlate respectively with systole and diastole of this multichambered heart. The periodic change in hemolymph direction is referred to as cardiac reversal. Intracellular recordings from muscles of the first abdominal cardiac chamber, the conical chamber, revealed pacemaker action potentials and the excitatory effect of local glutamate application, which initiated retrograde contractions in semi-intact preparations. Unilateral electrical stimulation of the transverse nerve containing the glutamatergic neuron that serves the conical chamber caused a chronotropic effect and initiation of retrograde contractions. This effect is distinct from that of peripheral crustacean cardioactive peptide (CCAP) neurons, which potentiate the anterograde beat. Cardiac reversal was evoked pharmacologically by sequentially applying CCAP and glutamate to the heart.

Illumination controls differentiation of dopamine neurons regulating behaviour

Specification of the appropriate neurotransmitter is a crucial step in neuronal differentiation because it enables signalling among populations of neurons. Experimental manipulations demonstrate that both autonomous and activity-dependent genetic programs contribute to this process during development, but whether natural environmental stimuli specify transmitter expression in a neuronal population is unknown. We investigated neurons of the ventral suprachiasmatic nucleus that regulate neuroendocrine pituitary function in response to light in teleosts, amphibia and primates. Here we show that altering light exposure, which changes the sensory input to the circuit controlling adaptation of skin pigmentation to background, changes the number of neurons expressing dopamine in larvae of the amphibian Xenopus laevis in a circuit-specific and activity-dependent manner. Neurons newly expressing dopamine then regulate changes in camouflage colouration in response to illumination. Thus, physiological activity alters the numbers of behaviourally relevant amine-transmitter-expressing neurons in the brain at postembryonic stages of development. The results may be pertinent to changes in cognitive states that are regulated by biogenic amines.

Innervation of the heart of the adult fruit fly, Drosophila melanogaster

The innervation of the adult abdominal heart of Drosophila melanogaster was studied by neuronal staining with green fluorescent protein and immunocytochemical techniques. The investigation was undertaken to determine whether the adult heart receives neuronal input or whether its complex activity must be considered independent from the nervous system. The larval heart lacks innervation, suggesting that the cardiac impulse is totally myogenic. At metamorphosis, segmental neural processes grow onto the myocardium. A pair of transverse nerves innervates bilaterally each cardiac chamber and its alary muscles. These nerve terminals are immunoreactive to glutamate and form unique synaptic structures on the ventral layer of longitudinal cardiac muscles of the conical chamber. This characteristic cardiac synapse may represent part of the neural mechanism controlling the retrograde heartbeat, and, thus, the cardiac reversal that is characteristic of adults. In addition, crustacean cardioactive peptide–immunoreactive fibers originating from peripheral, bipolar neurons (BpNs) fasciculate with the transverse nerve projections and terminate segmentally throughout the abdominal heart. An additional cluster composed of four large, CCAP‐positive neurons innervates the terminal chamber. The cardioacceleratory effect of CCAP release at this location may modulate the properties of a pacemaker producing the anterograde heartbeat. J. Comp. Neurol. 465:560–578, 2003.

Neuronal control of heart reversal in the hawkmoth Manduca sexta

Abstract. Cardiograms demonstrate that heart activity of Manduca sexta changes from larva, to pupa, to adult. The larval heart has only anterograde contractions. During metamorphosis, heart activity becomes a cyclic alternation of anterograde and retrograde contractions. Thus, the adult heart has both an anterograde and a retrograde pacemaker. External stimuli also can initiate cardiac reversal. Cardiac reversal is blocked by tetrodotoxin, indicating that reversal is under neuronal control. A branch of each dorsal nerve 8 innervates the posterior chamber of the heart, the location of the anterograde pacemaker. Only retrograde contractions occur when dorsal nerves 8 are cut. Stimulation of ml–1 8 initiates anterograde contractions; when stimulation ceases, the heart reverses to retrograde contractions. These experiments indicate that the anterograde pacemaker receives neural input that makes it the dominant pacemaker. In the absence of neural input this pacemaker is inactive, and the retrograde pacemaker becomes active. Application of crustacean cardioactive peptide accelerates the heart but does not eliminate cardiac reversal. The terminal chamber of the heart is also innervated by a branch of each dorsal nerve 7; stimulation of this nerve increases the strength of contraction of the terminal chamber but has no effect on contractions of the remainder of the heart or on cardiac reversal.

Innervation of the Heart and Aorta of Manduca sexta

Innervation of the heart and aorta of Manduca sexta was studied by using anatomic, neuronal tracing and immunocytochemical techniques. The study was undertaken to provide a foundation for investigating the neural mechanisms controlling cardiac reversal in adults. Lateral cardiac nerves were not found in the larval or adult heart. The larval heart and aorta seem to lack innervation, but a neurohemal system for the release of a cardioactive peptide is associated with the larval alary muscles. At adult metamorphosis, this neurohemal system regresses, and, at the same time, processes grow onto the anterior aorta. These processes seem to be neurohemal and originate from two pairs of neurosecretory cells located in the subesophageal ganglion. This system is immunoreactive to cardioactive peptides and may function, therefore, in hormonal modulation of the activity of the adult heart. Also during metamorphosis, synaptic innervation develops on the terminal heart chamber, and this innervation is from axons extending through the seventh and eighth dorsal nerves of the terminal abdominal ganglion. These axons originate from cells that have been identified as serial homologs of motor neuron‐1 of other abdominal ganglia. These neurons are immunoreactive to a cardioactive peptide, and this peptide probably modulates the synaptic innervation of the terminal heart chamber. During metamorphosis, the target of the motor neurons‐1 of the seventh and eighth segments becomes respecified from larval skeletal muscles to the terminal chamber of the adult heart. J. Comp. Neurol. 4406:245–260, 2001.

Contexts for Dopamine Specification by Calcium Spike Activity in the CNS

Calcium-dependent electrical activity plays a significant role in neurotransmitter specification at early stages of development. To test the hypothesis that activity-dependent differentiation depends on molecular context, we investigated the development of dopaminergic neurons in the CNS of larval Xenopus laevis. We find that different dopaminergic nuclei respond to manipulation of this early electrical activity by ion channel misexpression with different increases and decreases in numbers of dopaminergic neurons. Focusing on the ventral suprachiasmatic nucleus and the spinal cord to gain insight into these differences, we identify distinct subpopulations of neurons that express characteristic combinations of GABA and neuropeptide Y as cotransmitters and Lim1,2 and Nurr1 transcription factors. We demonstrate that the developmental state of neurons identified by their spatial location and expression of these molecular markers is correlated with characteristic spontaneous calcium spike activity. Different subpopulations of dopaminergic neurons respond differently to manipulation of this early electrical activity. Moreover, retinohypothalamic circuit activation of the ventral suprachiasmatic nucleus recruits expression of dopamine selectively in reserve pool neurons that already express GABA and neuropeptide Y. The results are consistent with the hypothesis that spontaneously active neurons expressing GABA are most susceptible to activity-dependent expression of dopamine in both the spinal cord and brain. Because loss of dopaminergic neurons plays a role in neurological disorders such as Parkinsons disease, understanding how subpopulations of neurons become dopaminergic may lead to protocols for differentiation of neurons in vitro to replace those that have been lost in vivo.

Reserve pool neuron transmitter respecification: Novel neuroplasticity

The identity of the neurotransmitters expressed by neurons has been thought to be fixed and immutable, but recent studies demonstrate that changes in electrical activity can rapidly and reversibly reconfigure the transmitters and corresponding transmitter receptors that neurons express. Induction of transmitter expression can be achieved by selective activation of afferents recruited by a physiological range of sensory input. Strikingly, neurons acquiring an additional transmitter project to appropriate targets prior to transmitter respecification in some cases, indicating the presence of reserve pools of neurons that can boost circuit function. We discuss the evidence for such reserve pools, their likely locations and ways to test for their existence, and the potential clinical value of such circuit‐specific neurotransmitter respecification for treatments of neurological disorders.

Remodeling of a larval skeletal muscle motoneuron to drive the posterior cardiac pacemaker in the adult moth, Manduca sexta.

During postembryonic development, a larval skeletal muscle motoneuron, MN‐1 in abdominal segments 7 and 8, becomes respecified to innervate the terminal cardiac chamber of adult Manduca sexta. Neural tracing techniques and electrophysiology were used in this study to describe the anatomical and physiological remodeling of this identified motoneuron. During metamorphosis the MN‐1 in segments 7 and 8 undergoes dendritic reorganization. Long new dendrites extend anteriorly in the terminal ganglion neuropil. Intracellular and extracellular recordings showed that broader action potentials, increased firing rate, and development of a bursting activity pattern accompany MN‐1 respecification. Cardiac mechanograms showed that MN‐1 activity bursts always correlate with the anterograde cardiac beat. Bilateral MNs‐1 fire at similar times to activate and sustain the putative cardiac pacemaker activity of the terminal chamber synergistically. After remodeling, MN‐1 output could be influenced rapidly by sensory inputs during evoked cardiac reversal. The effect is exerted by inhibition of MN‐1 firing that, in turn, causes early blockade of the anterograde beat and reversal to the retrograde direction of beat. J. Comp. Neurol. 478:126–142, 2004.

HIV-1 TAT protein enhances sensitization to methamphetamine by affecting dopaminergic function

Methamphetamine abuse is common among humans with immunodeficiency virus (HIV). The HIV-1 regulatory protein TAT induces dysfunction of mesolimbic dopaminergic systems which may result in impaired reward processes and contribute to methamphetamine abuse. These studies investigated the impact of TAT expression on methamphetamine-induced locomotor sensitization, underlying changes in dopamine function and adenosine receptors in mesolimbic brain areas and neuroinflammation (microgliosis). Transgenic mice with doxycycline-induced TAT protein expression in the brain were tested for locomotor activity in response to repeated methamphetamine injections and methamphetamine challenge after a 7-day abstinence period. Dopamine function in the nucleus accumbens (Acb) was determined using high performance liquid chromatography. Expression of dopamine and/or adenosine A receptors (ADORA) in the Acb and caudate putamen (CPu) was assessed using RT-PCR and immunohistochemistry analyses. Microarrays with pathway analyses assessed dopamine and adenosine signaling in the CPu. Activity-dependent neurotransmitter switching of a reserve pool of non-dopaminergic neurons to a dopaminergic phenotype in the ventral tegmental area (VTA) was determined by immunohistochemistry and quantified with stereology. TAT expression enhanced methamphetamine-induced sensitization. TAT expression alone decreased striatal dopamine (D1, D2, D4, D5) and ADORA1A receptor expression, while increasing ADORA2A receptors expression. Moreover, TAT expression combined with methamphetamine exposure was associated with increased adenosine A receptors (ADORA1A) expression and increased recruitment of dopamine neurons in the VTA. TAT expression and methamphetamine exposure induced microglia activation with the largest effect after combined exposure. Our findings suggest that dopamine-adenosine receptor interactions and reserve pool neuronal recruitment may represent potential targets to develop new treatments for methamphetamine abuse in individuals with HIV.