Selective dependence of mammalian dorsal root ganglion neurons on nerve growth factor during embryonic development.

We have investigated the NGF dependence of dorsal root ganglion (DRG) neurons in mammals using a paradigm of multiple in utero injections of a high titer anti-NGF antiserum. We have determined the specificity of our antiserum in relation to other members of the NGF neurotrophin family and found no cross-reactivity with brain-derived neurotrophic factor (BDNF) or neurotrophin-3 (NT-3). To identify various classes of DRG neurons, we have stained their characteristic central projections with Dil. We show here that the NGF dependence of DRG neurons is strikingly selective. Although a majority of DRG neurons are lost after NGF deprivation during embryonic life, these are almost exclusively small diameter neurons that project to laminae I and II of the dorsal horn and presumably subserve nociception and thermoreception. Larger neurons that project to more ventral spinal laminae and subserve other sensory modalities do not require NGF for survival. These NGF-independent DRG neurons likely require one of the more recently identified neurotrophins, BDNF or NT-3.

Dorsal root ganglion neurons expressing trk are selectively sensitive to NGF deprivation in utero.

In utero immune deprivation of the neurotrophic molecule nerve growth factor (NGF) results in the death of most, but not all, mammalian dorsal root ganglion (DRG) neurons. The recent identification of trk, trkB, and trkC as the putative high affinity receptors for NGF, brain-derived neurotrophic factor, and neurotrophin-3, respectively, has allowed an examination of whether their expression by DRG neurons correlates with differential sensitivity to immune deprivation of NGF. In situ hybridization demonstrates that virtually all neurons expressing trk are lost during in utero NGF deprivation. Most, if not all, neurons expressing trkB and trkC survive this treatment. In contrast, the low affinity NGF receptor, p75NGFR, is expressed in both NGF deprivation-resistant and -sensitive neurons. These experiments show that DRG neurons expressing trk require NGF for survival. Furthermore, at least some of the DRG neurons that do not require NGF express the high affinity receptor for another neurotrophin. Finally, these experiments provide evidence that trk, and not p75NGFR, is the primary effector of NGF action in vivo.

Administration or deprivation of nerve growth factor during development permanently alters neuronal geometry.

We investigated whether the administration or deprivation of a neuronal growth factor during development can permanently alter the dendritic architecture of sensitive neurons. Nerve growth factor (NGF) or NGF antiserum treatment in the first 2-3 postnatal weeks markedly affected the survival, size, and dendritic arborization of mouse sympathetic ganglion cells acutely. Six months after the completion of treatment, the number of surviving neurons, cell body size, and higher order dendritic branching had changed considerably from their values at 3 weeks, suggesting that these parameters remain malleable throughout postnatal life. However, the number of primary dendrites, a fundamental determinant of organization within sympathetic ganglia, was permanently altered by the neonatal treatment protocol. The idea emerging from this study is that NGF influences the elaboration of primary dendrites by sympathetic ganglion cells only during a critical developmental period. In maturity, NGF acts as a "maintenance" factor necessary for normal neuronal function and survival, but neurons lose the capacity to respond with wholesale rearrangements of dendritic architecture.

Cardiovascular and respiratory responses to electrical and chemical stimulation of the hippocampus in anesthetized and awake rats.

Electrical (30-60-s trains of 0.25-ms pulses at 25 Hz, currents 10-150 microA) and chemical (microinjections of 0.1-0.5 microliters of a 1.0 M glutamate solution) stimulation of the hippocampal formation in the anesthetized and the awake rat evokes marked decreases in heart rate, blood pressure and slower, deeper, more regular respirations. Artificial ventilation (2 ml/breath; 100 breaths/min) has no effect on the cardiovascular responses, indicating that these effects are not secondary to respiratory changes. Administration of methyl atropine (0.4 mg/kg) eliminates the bradycardia response and attenuates or obliterates the blood pressure response but does not alter the respiratory response. This suggests that the cardiovascular responses are mediated partially by the vagus nerve and partially by sympathetic influences. Ablation of the medial frontal cortex, a visceral motor region which projects directly to the nucleus of the solitary tract and which receives a heavy direct projection from the CA1 and subicular cell fields of the ventral hippocampus, markedly attenuates or eliminates the cardiovascular and respiratory responses to stimulation of the ventral but not the dorsal hippocampus. The possibility that the medial frontal cortex may be a relay by which the hippocampus influences cardiovascular responses, including those observed during stress, is discussed.

Hippocampal input to a "visceral motor" corticobulbar pathway: an anatomical and electrophysiological study in the rat.

The hippocampus has previously been shown to influence cardiovascular function, and this effect appears to be mediated by the connection the hippocampus has with the infralimbic area of the medial frontal cortex (MFC), a region which projects directly to the nucleus of the solitary tract (NTS) in the dorsal medulla. In the present study, anatomical and electrophysiological techniques were utilized to determine the degree of convergence of hippocampal input to the MFC on neurons in the MFC which project to the NTS. Injections of the anterograde and retrograde neuroanatomical tracer wheat-germ agglutinin-horseradish peroxidase (WGA-HRP) into the NTS retrogradely labelled cells in the infralimbic and prelimbic regions of the MFC. Injections of WGA-HRP into the ventral hippocampus anterogradely labelled terminals in the MFC which, at the light microscopic level, closely overlapped the origin of the descending projection from the MFC to the brainstem. Electron microscopic analysis revealed that anterogradely labelled terminals make synaptic contact primarily on dendritic processes in the neuropil adjacent to retrogradely labelled cells. In addition, anterogradely labelled terminals did, in some cases, make synaptic contact on the somas of retrogradely labelled cells. Electrical stimulation of the NTS antidromically activated cells in the infralimbic and prelimbic areas of the MFC. The average latency of antidromic activation was 30 msec, corresponding to a conduction velocity of approximately 0.7 m/s. Electrical stimulation of the ventral hippocampus orthodromically activated cells in the MFC. With an appropriate delay between the hippocampal and NTS stimuli, the orthodromic and antidromic potentials could be made to collide. The results of this study establish a structural as well as functional link between the hippocampus and NTS-projection neurons in the MFC.

Pathologic alterations in pre- and postsynaptic elements in aged mouse sympathetic ganglia.

Dysfunction of the sympathetic autonomic nervous system is an increasingly recognized, although poorly understood, complication of increasing age in experimental animals and man. In this study of young adult (4-6 months old) and aged (12-24 months old) mice we have examined the ultrastructural appearance of perikarya, dendritic processes, preterminal axons, and synapses in selected sympathetic ganglia as well as the three-dimensional structure of the dendritic arborizations of principal sympathetic neurons using intracellular injections of Lucifer Yellow. Ultrastructural examination demonstrated numerous markedly enlarged presynaptic terminal axons and synapses which distorted the contours of perikarya and dendrites of neurons within the prevertebral celiac/superior mesenteric and paravertebral superior cervical and stellate sympathetic ganglia of aged mice. Dilated preterminal axons had the distinctive ultrastructural appearance of neuroaxonal dystrophy, a pathologic process described in a wide variety of clinical and experimental entities. Dystrophic axons were identical in ultrastructural appearance in young and old animals, differing only in frequency. A distinctive type of ultrastructural alteration, characterized by markedly distended neurites containing numerous vacuoles, was confined to the superior cervical ganglia and also increased in frequency with aging. Although many intraganglionic vacuolated processes disappeared with surgical interruption of the cervical sympathetic trunk, which contains the preganglionic axons innervating the superior cervical ganglia, others persisted. In addition, the presence in some processes of admixed ribosomes, lipofuscin, or continuity with the cell body indicated that numerous neuritic alterations within aged sympathetic ganglia were likely of dendritic origin. Intracellular injections of Lucifer Yellow into principal sympathetic neurons demonstrated that the dendritic arborizations of the celiac/superior mesenteric ganglia neurons of young adult mice were significantly more complex and extensive than those of the superior cervical ganglia. Sympathetic neurons of aged superior cervical ganglia, but not superior mesenteric ganglia, appeared significantly smaller with regard to total dendritic length, extent, and branching when compared to those of young animals. In the aged superior cervical ganglia, short, stunted dendritic processes also exhibited large, focal, often multiple, swellings, a phenomenon infrequently observed in the superior cervical ganglia of young animals. The celiac/superior mesenteric ganglia of aged or young adult mouse failed to exhibit comparable dendritic swellings.

Nerve growth factor regulates sympathetic ganglion cell morphology and survival in the adult mouse.

We have investigated the effects of prolonged systemic injections of nerve growth factor (NGF) and its antiserum on the survival and morphology of sympathetic ganglion cells in adult mice. Using intracellular injections of Lucifer yellow in lightly fixed superior cervical ganglia, we show that total dendritic lengths of ganglion cells are increased 29% after 2 weeks of NGF treatment. The increased dendritic length is characterized by increased branching within the dendritic arborization and not by the addition of new primary dendrites. In addition, cell soma cross-sectional area was increased 45%. Conversely, administration of NGF antiserum for 1 month decreased total dendritic length by 33%, decreased ganglion cell body size by 26%, and reduced the number of neurons in the ganglion by 25%. After 3 months of NGF antiserum treatment, the number of neurons in the ganglion was reduced a total of 41%. NGF antiserum treatment for 1 month in aged (22 months old) animals reduced ganglion cell body size by 21% and cell number by 22%, decreases that are comparable to those observed in young adult animals. Our results indicate that, even in maturity, sympathetic ganglion cells remain dependent on NGF for survival and maintenance of dendritic geometry, and this dependence continues into old age.

PDGF A-chain gene is expressed by mammalian neurons during development and in maturity.

Platelet-derived growth factor (PDGF) may be a critical factor in the temporal differentiation of glial elements in the mammalian central nervous system. We have used in situ hybridization and immunoperoxidase staining to investigate the localization of PDGF A and have observed high levels of PDGF A-chain mRNA and immunoreactive PDGF A in neurons of embryonic and adult mice. PDGF A-chain expression was shown to be developmentally regulated and tissue specific. Every neuronal population examined in the central and peripheral nervous systems expresses PDGF A transcripts. Variable, significantly weaker signals are observed in glial cells. In contrast to known neurotrophic factors, the PDGF A transcripts are widely distributed among neurons. This generalized distribution of PDGF A transcripts, together with the known effects of PDGF on glial cells in vitro, suggests a unique role of neurons in regulating the proliferation and differentiation of glial cells in vivo.

Endocrine physiology in a patient-centered learning curriculum.

The medical curriculum at the University of North Dakota School of Medicine and Health Sciences has recently been redesigned into a problem-based/traditional hybrid model that utilizes an integrated organ systems-based approach to teach basic and clinical sciences. The number of lecture hours in general has been greatly reduced, and, in particular, lecture hours in physiology have been reduced by 65%. Students learn basic science in small groups led by a faculty facilitator, and students are responsible for a great deal of their own teaching and learning. The curriculum is centered around patient cases and is called patient-centered learning (PCL). The curriculum includes traditional lectures and laboratories supporting faculty-generated learning objectives. Endocrine physiology is taught in year one, utilizing four weeks of patient cases that emphasize normal structure and function of endocrine systems. Endocrine physiology is revisited in year two, which is primarily focused on pathobiology. The PCL curriculum, with emphasis on the endocrine component, is described in detail along with key portions of an endocrine case.